AAMC FUNCTIONAL CONCEPT 1: BIOMOLECULES = STRUCTURE & FUNCTION

1A. STRUCTURE & FUNCTION OF PROTEINS & THEIR CONSTITUENT AAs

BUILDING BLOCKS

Building Block Polymerizes to form…. Chemical bonds Macromolecules

Monomers  Dimer Covalent bonds * Polymer
 Trimer
 Tetramer
 Oligomers
Amino acids  Dipeptide Peptide bonds  Polypeptide
 Tripeptide  Protein
 Tetra/oligopeptide
Monosaccharides  Disaccharide Glycosidic bonds  Polysaccharide
(“simple sugars” **)  Tri/Tetra/Oligosaccharide
Nucleotides  Nucleotide dimer Phosphodiester bonds  Polynucleotides
 Tri/Tetra/Oligomer  Nucleic acids
** Exceptions  certain circumstances polypeptides are considered monomers & may bond
non-covalently to form dimers (i.e. higher orders of protein structure)

** Disaccharides  sugars
- Sucrose = glucose-fructose dimer = ‘table sugar’
- Lactose = glucose-galactose = ‘milk sugar’

 PROTEINS

 Functions:
o Act as catalyst  speeding up biochemical reactions used by cells
 Ex: Production of ATP involves enzymes (proteins – biological catalyst)
o Transport
 Ex: Hb  protein that transports O2 from lungs into tissues and cells
 Embedded inside cell membranes  they act as transport proteins and
allow movement of molecules and ions across cell membrane of cells
o Structure
o Mobility
 Ex: Sperm cells  flagellum (movement)  composed of different types
of proteins
o Protection/Immunity
 Ex: against pathogens  Ab and Ag consist of proteins
o Communication
 Ex: Hormones (peptide hormones) involved in intracellular and cell to cell
communication
  LINEAR POLYMERS  Consist of building blocks = AMINO ACIDS (20)
o Proteins are polymers of amino acids
 Peptides = aa subunits = residues
 DIpeptide = 2 aa residues
 TRIpeptide = 3 aa residues
 OLIgopeptide = up to about 20 residues
 POLYpeptides = more than 20 residues  longer chains

 DIFFERENCE B/W POLYPEPTIDE & PROTEIN

Polypeptide Protein
 Polymer  simple chain AAs  Complex  folded polypeptides
 AAs linked w/ covalent peptide  Noncovalent weak bonds b/w folding
bonds polypeptides
o H bonds
o Ionic bonds
o van der Waal bonds
 Characterizes primary structure  Can exist as secondary, tertiary, OR
quaternary structure
 NO functional properties  due to  Functionally complex & active  w/
simplet structure specific ligand-binding sites formed
on surface by folding of polypeptide
chains

 Polypeptides & Proteins

 Natural & essential organic compounds of cell
 BOTH composed of AAs
o AAs held together by:
 covalent bonds = peptide bonds (amide bond)  form polypeptide 
residues in peptides are joined together by peptide bonds
o AAs link together to form:
 Peptides
 Polypeptides
 Proteins
o PEPTIDE BOND forms b/w  COO- group of 1 aa & NH+3 group of another aa
 forms functional group –C(O)NH--
o How many possible sequences are there if we have these 10 amino acids and
each one of these amino acids can be anyone of 20 amino acids? (use
probability)
 Aa #1 = 20 possibilities
 Aa #2 = 20 possibilities
 Multiple these out to find the total possibilities that we can have
 2010 = 1.024 x 1013 possibilities of a protein that consists of 10 aa
 Means there is a great # of different types of sequences of the proteins
o Sequence of aa importance  determines its 3D structure
  Linear sequence of aa can fold into 3D structure spontaneously
 Once proteins form their 3D structure  their function is determine
 3D structure of protein = determines proteins function

o What is the difference between one aa and another aa?

 AA’s differ  based on the side chain group (functional group)
 Glycine = H
 Cysteine = SH
  Form disulfide bond linkages  OXIDATION of -SH
groups of 2 cysteines allows them to participate in covalent cross-
links
o = DISULFIDE BONDS (-S-S)  & form CYSTINE
DIMER
 DISULFIDE bonds b/w 2 cysteines can occur:
o W/in
OR
o B/w
 Stabilizes 3D structure
 Different functional groups have different capabilities and arrangement
and sequences of these amino acid functional groups determines how the
proteins are involved in helping the different reactions inside our body
 Imp. With biological catalyst  because these catalyst use these
functional groups to carry out different types of reactions in which
they speed up the processes inside our body

 AAs STRUCTURE
 AAs R group makes it:
 ASYMMETRIC
 Allows it to exist in 2 mirror-image forms (D & L)  STERIOISOMERS
 ONLY L = mammalian proteins

 Absolute configuration @ ALPHA position

 D&L
o = RELATIVE configuration  relative relationship b/w AAs and D & L
sugars – small sugar -glyceraldehyde
 R&S
o = ABSOLUTE configuration  tells absolute order of priority groups
  D & L AND R & S = BOTH designate ENANTIONMERS
o D & L = different from R & S
  L NOT always S & D NOT always R

 Alpha AA = Central Carbon

o = contains amino group and acid group separated by ONLY 1C atom

 AAs as DIPOLAR IONS

CLASSIFICATION
 @ LOW pH = Cation (AA)
 @ HIGH pH = Anion (AA)
 @ pH = pI = Zwitterion (AA)
 Neutral

 AA CLASSIFICATION
 HYDROPHOBIC = R groups DOES NOT contain any stuff
 HYDROPHILIC = R groups contain acids, bases, amines, OR alcohols
 Rigid Proteins
o Involved in giving our cells our structure
 Ex: cytoskeleton
 Non-Rigid Proteins
 o Involved in processes that involve flexibility

 Proteins don’t act by themselves  interact with other proteins or other macromolecules
to form protein complexes and carry out a function  that create processes and rxns
o Ex: DNA replication  not just one protein involved in DNA replication large
protein complexes required to create new DNA
o Ex: transport proteins in cell membrane (phospholipids)  interact w/
phospholipids to transport molecules and ions across the cell membrane

 Proteins are encoded by the DNA in our body

o DNA  RNA  protein

 Proteins are built from amino acids

o Alpha amino acids

 Absolute configuration
o Determine by prioritize different types of atoms attached to the center chiral
carbon
o Give 4 groups value that ranges b/w 1-4
o 1 = atom/group that contains highest atomic #
o Orient molecule so H group points into board so we cannot see it and 3 groups
point out
o ** majority of L – amino acid isomers (w/ the exception of 2) have the S
absolute configuration = counterclockwise

 PROTONATION and DEPROTONATION:

o pH of solution in which aa is in determines whether or not the amino group or
carboxylic acid group are protonated or deprotonated  adding neutral pH = 7
(around this) aa exist predominantly in dipolar form = zwitterion form pH = 7
 = amino group = protonated = + charge
 carboxylic acid group = deprotonated = - charge
  have dipolar species = have 2 dipole moments

o Decrease pH = more acidic

 ACIDIC pH = 1  so many H+ ions in solution the COO- = protonated =
overall charge = +1 = positively charged aa form
 Amino acids exist in positively charged form
 As begin to increase pH  decrease [H+ ions] inside solution  @
around 2, H on the carboxylic acid group deprotonates (loses H atom) and
gains full negative charge
o Increase pH = more basic
 pH = 4,5,6, and 7 = zwitterion ion form
 continue increasing pH and decreasing H+ ions in solution  little H+
ions amino group H deprotonates to increase the amount of H+ ions in
solution
  pH = 9  begins here  amino group deprotonated  negatively charged
aa form

 **Favorite question of examiners:

o AA w/ more than 1 amine group:
 Question asks to draw structure in “excess HCl”  PROTONATE
BOTH NH2 groups

protonate both NH2 groups

PROTEIN STRUCTURE
 Folded polypeptide  hydrophobic core & hydrophilic surface
 Protein folding and stabilization depend on noncovalent forces
 In PROTEIN = ONLY N-term -NH3+ & C-term -COO- & any ionized AA R-group =
CHARGED

http://studyhall.leah4sci.com/wp-
content/uploads/2018/03/Leah4scimcatcheatsheetcollection2017.pdf

PEPTIDE BOND = amide bond b/w 2 aa

 Rigid bond
 Trans configuration
 Uncharged BUT polar
o Polar = cuz participate in H bonding  i.e. α-helix secondary structure
 Proteins are formed by the folding of polypeptide chains
 Polypeptide chains are formed by linking amino acids together  links are called
peptide bonds
 1) form peptide bond b/w 1st two amino acids
 Peptide bonds are formed by nucleophilic addition-elimination reaction b/w carboxyl
group of one aa and amino group of another aa
o Give off H2O during process
 Resonance delocalization of electrons has a lot of double bond character
o Rigid  not much rotation
o Planar
o Still have rotation of alpha carbons
 Backbone of chain  forms pattern of atoms
o N  alpha carbon  carbonyl carbon  N  alpha carbon  carbonyl carbon
 -C=O & -NH groups of peptide bonds of polypeptide = NET neutral charge
 o N C  C N C C  N
o Always start with N and always end with carbonyl carbon
o Each aa = residue
 Breaking this bond by  hydrolysis = form 2 free aa
o N-C = peptide bond
o Two common means
 1) strong acids = acid hydrolysis
 When combined w/ heat = non-specific way of cleaving peptide
bonds
 End up w/ jumble of aa
 2) proteolytic enzymes
 Specific cleavage/breaking of peptide bond w/ help of protease
 Can chose which peptides bonds you cleave = cleave b/w certain
specific aa
 Ex: Trypsin = only cleaves on carboxyl side of basic aa Arginine
and Lysine
o same enzyme produced by pancreas to help us digest food
o N-terminal – Thr – Arg – His – Pro – Lys – Val – C-
terminal

 cleave where red arrow is when using

trypsin

 End up with 3 different fragments after the addition of

trypsin since it cleaves in these specific
places
 Formation of peptide bond
 Example of condensation or dehydration rxn  due to removal of H2O molecule OR a
acyl substitution rxn  which can occur w/ all carboxylic acid derivatives

 Proline & Glycine are grouped together because they BOTH play a role in disrupting pattern
found in secondary protein structure = alpha helix
 Proline & Glycine are grouped together because they BOTH play a role in disrupting a
particular pattern found in secondary protein structure = alpha helix
 Proline  induces kinks into the alpha helix
 Glycine  so flexible around alpha carbon it induces kinks into alpha helix

 Hb  found in RBCs  each RBC full of Hb protein

 Hb protein = responsible for picking up O2 when RBC flows through vessels of lungs
and transports O2 to tissues (tissues = groups of similar cells)  each of cells in tissues
uses it to make ATP = energy source for metabolic processes of cells
 Hb = think of as a car
  O2 = passenger of car
 Amino acids = parts that come together to form the car (proteins)
 Amino acids = building blocks of Hb protein
 Without amino acids this process could NOT occur
 Chiral carbon = 4 unique groups bound to it  optical activity  if shoot plain
polarized light at aa  because chiral it would rotate toward that light
 Amino group
 Carboxylic acid group
 H group
 R group = side chain
 L-amino acid = ONLY type you found within humans

 ISOELECTRIC POINT (PI)

 Molecule (amino acid) exists in NEUTRAL form for 0 charge
 Know this for amino acid because can predict if that amino acid is charged at a certain
pH
 ACIDIC AAs & proteins w/ lots of acidic side chains = LOWER isoelectric point
 BASIC AAs & proteins w/ lots of basic side chains = HIGHER isoelectric point
 Change in amino acid is happening @ each pka value

 ELECTROPHORESIS
 Protein = charged
 Electric field forces protein to travel through gel
 LARGER charge = MORE electrical force = travels FASTER
 SMALLER protein = squeezes through easier = travels FASTER
 Structure NO role  because SDS usually added  denature protein

 NON-ENZYMATIC PROTEIN FUNCTION

 Binding:
 Active site BINDS substrate
 STRONGER binding = LOWER Kd value
 Stronger binding DOES NOT = MORE efficient enzyme  if it binds substrate and
NOT let go, then CANNOT catalyze new substrate
 STONGER binding = BETTER Ab

 CLASSIFICATION OF AMINO ACIDS

MNEUMONIC: MNEUMONIC:
AROMATIC SIDE CHAIN AAs = HTTP COOH & CONH2-CONTAINING / AMIDE
SIDE-GROUPS AAs = Ag
Histidine
Tryptophan Aspartate Asparagine = N = Asn
Tyrosine Glutamate Glutamine = Q = Gln
Phenylalanine

MNEUMONIC:
OH-CONTAINING SIDE-GROUPS AAs – can
be phosphorylated
= SHOTT
 MNEUMONIC:
ACIDIC – POLAR (-) AAs – excitatory
brain NTs-make post-synaptic neurons
MORE likely to fire

AS Peter Digested the GLUE

ASpartate = D
GLUtamate = E
MENUMONIC: = AAGG
BASIC – POLAR (+) AAs
= HALeluijah Aspartate
Asparagine
Histidine Glutamate
Arginine Glutamine
Lysine

BASICally HIS Lost Kid Always Returns

H =Histidine
MNEUMONIC:
K = Lysine
NON-POLAR-HYDROPHOBIC AAs
R = Arginine
= GLaciers in ALAska VALIantly Locate ISOlated PROwlers MNEUMONIC:
AROMATIC AAs
GLycine = PfTTP – since they have a
ALAnine smell
VALIne
Locate Phenylalanine
ISOleucine Tyrosine
PROline Tryptophan

MNEUMONIC:
S-CONTAINING SIDE-GROUPS AAs – each can link to other sulfur containing AAs
through oxidation of their sulfhydryl bonds to form S-S BONDS

= SCUM

(S) = Sulfur
Cysteine
(U)
Methionine

 PROPERTIES OF AAs
  1) Acid & Basic AAs = CHARGED AAs
 2) ALL aliphatic AAs + methionine  Sulfur-containing AA – and phenylalanine &
tryptophan = aromatic AAs = NONPOLAR AAs
 3) ALL alcohols & amides + cysteine, which is Sulfur-containing AA, & tyrosine =
aromatic AA = POLAR AAs

 AAs
 Similar in the chemical properties of the side chain  depends on side chain….
 charge
 H bonding ability
 acidic vs. basic
 Alkyl Amino Acids = nonpolar, hydrophobic, non-reactive
 Glycine
 Alanine
 Valine
 Methionine
 Leucine
 Isoleucine
 Proline
 Exception: side chain forms ring structure with amino group and backbone of
molecule
 Aromatic Amino Acids = nonpolar, hydrophobic
 Made up of Carbons and Hydrogens
 3 RULE OF AROMATICITY
 1) Ring structure
  2) MUST be planar  w/ unhybridized P orbitals overlapping to form a
CONTINUOUS ring of planar orbitals
 3) Ring MUST follow HUCKEL’S RULE  have 4n + 2 electrons in its
system of conjugated P-orbital clouds
 n = integer
 Neutral Amino Acids: => Polar, Hydrophilic
 Side chain contains oxygen or sulfur atom  localized – charge over atom and +
charge over rest of side chain
 Not strongly polar enough to qualify as acid or base
 Acidic Amino Acids:
 Aspartic Acid
 Glutamic Acid
 Basic Amino Acids:
 Side chain contains nitrogen atom  nitrogen = proton acceptor = why these aa
qualify as basic
 Histidine
 Lysine
 Arginine

 HYDROPHOBIC SIDE CHAIN AMINO ACIDS

 Leucine & Isoleucine are more nonpolar than Valine
 Isoleucine contains chiral atom (mirror image) = only Nanta mero s isoleucine that
we find in our proteins
 H atom coming out of board
 Valine is more nonpolar/electronegative than Alanine
 more C-H atoms = more hydrophobicity w/in that aa
 Alanine (Ala, A)
 Valine (Val, V) - Each of these 4 aa have
 Leucine (Leu, L) hydro-carbon side chains
 Isoleucine (Ile, I) - C-H = nonpolar, non-reactive
 Methionine
 Side chain = Sulfur atom
 Electronegativity Carbon = 2.55
 Electronegativity Sulfur = 2.58
 These values are essentially the same  why these bonds are nonpolar and
side chain is nonpolar
- All 3 contain a ring
 Phenylalanine structure
 Benzene ring (C-H) = side chain - Still nonpolar and non-
 VERY HYDROPHOBIC and non-reactive reactive due to large ring
 Tyrosine structure
 Benzene ring w/ hydroxyl group = side chain
 Contains electronegative O and N
 LESS HYDROPHOBIC and less reactive
 Tryptophan
 Indole group = side chain
  Phenyolonine more hydrophobic than Tyrosine
 Nonpolar side chain groups tend to pack together/aggregate in the protein these are side
chains that point inward of the protein structure
 Inside our cells and bodies are made of polar water molecules and proteins are found
inside the polar molecules  hydrophobic side chains display the hydrophobic effect
and create this packed structure found inside that protein

POLAR AAs = MORE REACTIVE than nonpolar amino acids

NONPOLAR & HYDROPHOBIC AAs = LESS likely to be

catalytically important

NONPOLAR & HYDROPHOBIC AAs = MORE important in

protein-protein interactions & forming the protein core

 UNCHARGED, POLAR SIDE CHAINS

** HYDROPHILIC  due to ability to undergo HYDROGEN BONDING
** Side chains = attachment sites for other compounds
 Phosphate groups
 Oligosaccharides

 Serine
 Hydroxyl group
 O atom = gain partial (-) charge because O is more electronegative than H
 H atom = partial (+) charge
 = dipole moment  from + end to – end = uncharged polar side chain
 Interact w/ H2O molecules
 Side chain is MORE BASIC than cysteine  meaning it is LESS ACIDIC  less
likely to lose its proton
 Highly polar  participate in hydrogen bonding
 Threonine
 Carbon contains methyl group
 Contain chiral carbon  only Enantiomer found in body
 Electronegative atom  dipole moment  polar side chain
 Highly polar  participate in hydrogen bonding
 Asparagine
 Carboxy amide group = side chain
 Amide side chain  DO NOT gain or lose protons w/ changes in pH  they DO
NOT become charged
 Glutamine
 1 additional CH2 group compared to asparagine
 Terminal end of side chain have carboxy amide group
 Amide side chain  DO NOT gain or lose protons w/ changes in pH  they DO
NOT become charged
 Cysteine
  Thiol group (-SH) side chain
 Sulfur is larger and less electronegative than oxygen  S-H bond is weaker than
the O-H bond  this leaves the thiol group in cysteine prone to oxidation

 GLYCINE:
 No enantiomer  because of 2 identical H atoms attached to Carbon
 ACHIRAL  optically inactive
 Side chain:
 ONLY H  BUT acts similar to alkyl chain side group
 NOT chiral (handedness)
 No C-H (NO HYDROCARBON side chain)  so NOT hydrophobic  but
because of its small size, it can easily interact with other hydrophobic side chains and
also interact with hydrophilic side chain
 Function:
 Protein flexibility
 Location:
 Exterior of proteins in β-bends = flexibility
 Smallest/Simplest AA
 Fit into either hydrophobic or hydrophilic environments

 PROLINE: = IMINO  contains BOTH imine & carbonyl

 Hydrophobic side chain
 Structure:
 α-amino N (2° amino group)  form rigid ring structure = disrupts 2° structure
 Only side that connects to alpha carbon and N  form 5 membered rigid ring 
structurally restrictive  helps w/ role in determining structures in types of proteins

 Alkoxide ion = poor leaving group  conjugate base of alcohol, fairly weak acid  pKa
15-19

Hydrogen bonding = attracted to F, Polypeptides HYDROPHILIC properties

O, & N  HIGH electronegativity = due to strong electronegativity of O2 in
H2O that attracts H+’s

 CHARGED, POLAR, SIDE CHAINS

 ALL contain full charge on side chain groups @ normal physiological pH  this is
what makes them very highly hydrophilic
 BASIC AAs
 Side chains @ physiological pH =7 = bare full (+) charge
 Lysine & Arginine ACCEPT H+’s to their R-groups @ physiologic pH to form
fully ionized – positively-charged
  (-NH3+ , NH2+ Lysine & Arginine = ALWAYS +
 LYSINE charge on side chain groups @
physiological pH = 7
 4 carbon atoms w/ 2 H atoms on each carbon
  @ terminal end of side group have primary amino group  have N bond to single
C
 ALL aa @ normal physiological pH have full + charge on that N because to
deprotonate N have to increase pH  make it basic because pka value of N = 10
 Pka = if pH = to pka then that is point @ which ½ are deprotonated and ½
protonated
 < pka 10 @ physiological pH = 7  all lysine aa will be protonated
 ARGININE
 Guanidinium group  pka = 12.5  all side chain groups @ physiological pH =
protonated  full + charge
 + charge is delocalized among different atoms  stabilizes this molecule =
resonance stabilized
 HISTIDINE  + or neutral @ pH = 7
 WEAK BASE  BUT when incorporated in a protein = side chain =
positively-charged OR neutral depending on ionic environment
 Ex: Hb binding to O2
 Basic = full + charge on side chain group  sometimes be neutral @
physiological pH = 7
 MOST stable AA @ physiologic pH
 Can protonate & deprotonate @ NEUTRAL pH
 Histamine precursor
 Pka = 6  can exist in protonated or deprotonated state @ neutral pH  depends
on what local environment is around this amino acid
 Imidazole group  pka = 6.0  N will gain H+ and be protonated  gain +
charge  + charge can be delocalized gains H atom delocalized among 2 atoms
and will be + charge that is delocalized = resonance stabilized (like arginine)
because aromatic ring
 Contains lower pka and because can be protonated or deprotonated @ pH = 7 
common amino acid that exists in the active site of enzymes (where rxn takes
place)

 ACIDIC AAs
 ** MOST exist in deprotonated form
 Polypeptides w/ NET (-) charge = exhibit electrostatic repulsion
 Side chains @ physiological pH = full (–) charge
 Because pka value of side chains low < 7
 Carboxylate ion groups  full (-) charge  (-) charge delocalized among the 2
electronegative O atoms = stabilizing effect
 Pka = 4.1  < 4.1 these groups are protonated BUT > 4.1 these groups are
deprotonated
 pH = 7 exist in deprotonated state
 Aspartate  aa form @ pH = 7 = deprotonated form of aspartic acid
 1 less C
 < pH= 4.7 = protonated  no longer aspartate = aspartic acid

  Glutamate  aa form @ pH = 7 = deprotonated form of glutamic acid
 Extra C
 < pH = 4.7 = protonated  no longer glutamate = glutamic acid

 In AQUEOUS ENVIRONMENTS of cell  BOTH amino group AND carboxyl group

=> ionized under physiological conditions  this is why AA viewed as dipolar ions

CATIONIC @ LOW pH

ANIONIC @ HIGH pH

 Chemical composition of side chain determines the characteristics of AA

o Hydrophilic OR hydrophobic
o Basic OR acidic
 R groups  divided into 4 categories based on distinct chemical compositions:
o Acidic
 Side chain contains carboxylic acids
o Basic
 Side chain contains amines
o Polar

 IONIZABLE AAs (7/20) = @ certain pH values these side chain groups can exchange H
ions  donate and accept H atoms and gives them ability to participate in acid-base rxns and
form ionic bonds
 Every aa contains alpha-amino group & alpha-carbonyl group that can lose/gain a H+ atom
@ some particular pH
 If side chain groups can form ions  ions have full charges and charges can participate
in forming ionic bonds w/ other macromolecules  this is what makes these 7 aa reactive
and gives them ability to participate in types of biological rxns
 7 ionizable amino acids
 On top of each of these aa side chains containing ionizable side chain groups every
amino acids also contains ionizable alpha carbon groups and ionizable alpha amino
group
 Histidine
 Ring w/ 2 nitrogen atoms  BUT NOT considered aromatic
 Ring called = imidazole
 Cysteine
 Lysine
 Has terminal primary amino group
 Tyrosine
 Arginine
 Side chain contains 3 nitrogen atoms
 Positive charge is delocalized over all 3 nitrogen groups
 Aspartic Acid
  Glutamic Acid
 Side chain
 ALL 20 aa contain alpha carboxyl group and alpha amino group  @ specific pH ALL
aa can gain or lose H atoms  body is pH = 7 alpha carbonyl (-) charge and alpha amino
(+) charge = cancel each other out and that charge depends on that side chain group of
that aa

 How are aa held together inside a protein?

 Peptide bond (covalent bond OR amide bond) = holds aa together
 Reactant side: AA A (side chain = R1) & AA B (side chain = R2)
 Product: C = dipeptide = 2 aa connected by peptide bond (green)
 Whenever a chemical reaction takes place  # of atoms always conserved due to
conservation of mass
 QUESTION: If these products C & D are higher in energy than reactants A & B, why do
these bonds not spontaneously break inside body inside protein holding aa?
 Because of a high activation energy
 Reverse reaction: product  reactants = is thermodynamically favorable but not
kinetically favorable (have to input a lot of energy to overcome this reverse activation
barrier – we don’t have that much energy under normal conditions @ pH 7 and
normal body temp. to overcome or for rxn to take place)
 To break peptide bond inside body have to use enzymes that decrease activation
barrier
 Activation energy  barrier energy needed that must overcome
 If the dipeptide formation is energetically unfavorable why doesn’t the reverse rxn take
place spontaneously?
 Activation energy is too high
 @ conditions of body temp. and pH 7 this rxn does not take place
 Peptide bond 3 facts
 1) form via dehydrolysis rxn  @ end of forming single peptide bond ALWAYS
release H2O
 Breaking of peptide bond is hydrolysis rxn  have to use H2O to break peptide
bond to give back the C its O and N its 2 H atoms
 2) peptide formation (biosynthesis of peptide bond) is thermodynamically
unfavorable process  ALWAYS have to input energy  have to take energy away
from ATP to form peptide bonds that hold aa together inside proteins
 3) reason that inside our cells these proteins & peptides DO NOT spontaneously
break apart because the reverse rxn is kinetically unfavorable and the forward rxn is
kinetically favorable
 Too much energy in the activation barrier for us to go in reverse from the
products  reactants
 Would have to increase temp. to high temp. OR use enzyme that decreases
activation energy and break peptide bond and go back to individual amino acids
 Peptide bonds = kinetically stable
 Forward rxn = kinetically favorable (stable)
 Reverse rxn = kinetically unfavorable (unstable)
 Alzheimer’s Dz
 Proteins in neurons become misfolded and form aggregates outside of neurons = amyloid
= clumps of misfolded proteins
 As amyloid builds up it interferes with brains ability to send messages  leads to
dementia and memory loss
 How proteins become properly folded?
 Proteins consist of 1 or more polypeptides
 (polypeptide/protein (interchangeable)

PROTEIN STRUCTURE: 4 LEVELS

 Primary structure = linear sequence  N to C (read)

 Determined by peptide bond linking each aa
 Describes the specific sequence of aa w/in protein w/in our polypeptide
 AA specific sequence in polypeptides  determines the final 3D conformation
of the polypeptide
 Determines final structure
 specific sequence of aa in polypeptide that determines the final 3D structure of
polypeptide
 aa’s in a polypeptide = aka residue
 ** have n # of aa = always have n-1 # of peptide bonds
 Ex: have 5 aa & 4 peptide bonds
 @ pH 7 = polypeptide ALWAYS has polarity  alpha-amino = full (+) charge &
alpha carboxyl group = full (-) charge = polarity due to 2 different charges
 Beginning of polypeptide ALWAYS @ (+) amino end
 End of polypeptide ALWAYS @ (-) carboxyl end
 ALWAYS read NC
 Repeating unit of 3 atoms = NCCNCCNCC (continue if keep
adding aa)  repeating section of polypeptide = backbone
 Variable groups of aa = side chains
 Polypeptide as whole has potential to form H bonds w/ other molecules  H bonds
formed as result of atoms in polypeptide creates the secondary structure
 Polypeptide chain has ability to form many hydrogen bonds 
 Because each 1 of these aa contains 2 types of groups  important in
secondary structure of proteins
 1) hydrogen bond donor = N-H group
 N = more electronegative than H and H has a partial (+) charge) 
ensures that if have another atom that is electronegative in close
proximity then the H will be donated to that electronegative atom
forming a H bond
 2) hydrogen-bond acceptor = C = O (carbonyl group)
 O more electronegative than C  O develops partially (–) charge and
if H w/ partially (+) charge is found in close proximity on the other
side of partially (-) O forming a H bond
 Nature of peptide bonds
 Peptide bonds are resonance stabilized  which means they have double
bond character  length b/w C=N somewhere b/w a single and double bond
 Double bond character  makes the peptide bond planar and prevents any
rotation among the peptide bond
 Single bond = can rotate
 Double bond = cannot rotate
 6 atoms that rely on the same plane  all molecules lie along the same plane
 Can draw 2 Lewis dot structures
 Actually structure is neither, it is somewhere in b/w
 We see predominantly the trans and not the cis
 2 alpha carbon groups point in opposite directions (never point in the same
directions)
 Only consider 2 aa that are connected by peptide bond (more of double
bond than single bond  so get cis/trans isomers)
 In all of peptide bonds have transconfiguration  alpha carbon points in
opposite direction, away from the other alpha carbon
 Trans = more thermodynamically stable & lower in energy
 Because in the trans case all the atoms point away  if they point
away there will no steric hinderance, no bumping of electrons or
atoms, and no electrostatic repulsion as result of bumping of atoms
 If flip it to cis isomer  now alpha carbons point along the same
direction, will be steric hinderance that increases the electrostatic
repulsion due to the charges on the atoms, drives the energy of cis
configuration to be higher than trans configuration
 In some cases do see cis configuration, but in majority of cases see
trans
 No rotation b/w atoms of peptide bonds
 Have linear polymer of aa but the final 3D structure of proteins is not
linear, it is convoluted  to go from primary to quaternary structure have
to rotate bonds but peptide bonds cannot rotate, so have other bonds that
rotate
 Torsion angles = angles that can rotate = allows sequence of aa to create
quaternary 3D structure of protein
 N-alphaC = torsion phi angle angle = rotation about the single bond of
the N & alphaC
 alphaC – torsion angle si C of carbonyl = rotation about the single
bond of the alphaC- C of carbonyl
 Why can these rotate and peptide bond can’t?
 Peptide bond = resonance stabilized and it is NOT a single bond 
it is mostly a double bond
 Torsion bonds are purely single bonds that can rotate opposed to
double bonds that cannot rotate
 ** due to steric hinderance and conclusions/bumping of near
atoms, NOT all of the phi and si rotations of possible because
some of those rotations will lead to steric hinderance
  Not all of these bonds can rotate in specific ways  good
thing: means that if there are a limited #’s of ways that our
rotations that these 2 torsion angles can take place, that means
the linear polymer of aa has to go through less number of these
rotations to end up with final 3D structure of protein
 Every protein contains a primary structure  unique aa unique
to that protein  sequence of aa determines what the 3D
structure of that protein is

 SECONDARY STRUCTURE:
 = repetitive motifs formed by backbone interactions
 Backbone interactions = H bonding b/w NH & C=O
 Way the linear sequence of aa fold upon itself
 Local interactions
 Spatial arrangement of those aa and that interaction of those aa that are found in close
proximity on that polypeptide chain
 Determined by backbone interactions held together by H-bonds
 Primarily  hydrogen bonds
 2 motifs OR patterns
 1) ALPHA HELIX  R groups sticking OUTWARD
 Right handed w/ 3.6 AAs per turn
 H bonds = WEAK but strongly collectively
 ALL but 1st & last peptide bonds are linked via INTRACHAIN H-
bonding
 H bonds form b/w peptide bonds 4 AAs apart & run parallel to helical axis
 Take polypeptide and wrap it around itself in coiled structure
 Hydrogen bonds run up & down stabilizing the coiled structure
 2) BETA-PLEATED SHEETS  R groups stick OUT above OR below sheet
 Stabilized by H-bonds
 Right handed twist in globular proteins
 Parallel vs. antiparallel sheets

 TERTIARY STRUCTURE:
 Higher order of folding w/in polypeptide chain
 Distant interactions
 Spatial arrangement of aa that are found far away from each other on that
polypeptide chain
 Caused electrostatic side chain – side chain interactions
 Many different folds that fold upon each other again
 Depends on distant group interactions
 Stabilized by hydrogen bonds
 Also have other interactions
 Van der Wall interactions
 Hydrophobic packing
  Have folded up protein found in H2O part of cell  H2O on exterior of
protein then find all of the polar groups on exterior interacting w/ H2O  on
interior find hydrophobic groups
 Disulfide bridge formation
 Interaction only b/w cystines  have thiol group on side group  Sulfur
atom can become oxidized and when oxidation occurs get the formation of
covalent bond b/w sulfur groups
 Location: formation on exterior of cell
 See formation of separate thiol groups on interior of cell
 Interior of cell has antioxidants that generate a reducing environment
 Exterior of cell lacks antioxidants = get oxidizing environment
 Which environment favors the formation of disulfide bridges?
 Extracellular space

 QUATERNARY STRUCTURE
 ONLY proteins w/ > 1 polypeptide chain  NOT ALL proteins have it
 ALL proteins have  primary, secondary, & tertiary
 = smaller globular peptides = SUBUNITS  functional protein form
 Separate chains/subunits joining TOGETHER
 Individual protein subunits = folded up proteins that come together to form complete
protein
 Bonding b/w multiple polypeptides
 EXAMPLES: BOTH have 4 subunits
 Hb
 Immunoglobulins
 Described by the interactions b/w these 4 polypeptides and w/in the completed
protein structure each individual polypeptide = subunits
 QUATERNARY STRUCTURE ROLES
 1) MORE stable by  further reducing surface area of protein complex
 2) REDUCE amt. of DNA needed to encode protein complex
 3) Bring catalytic sites CLOSE together  allowing intermediates from 1 rxn to
be DIRECTLY shuttled to 2nd rxn
 4) ** Induce COOPERATIVITY or ALLOSTERIC EFFECTS
 1 subunit undergoes conformational OR structural changes to  enhance OR
reduce activity of other subunits

 4 subunit protein = tetramer

 2 subunit protein = dimer
 3 subunit protein = trimer
 Above 4 subunit protein = multimer
 Completely, properly folded up protein = proper conformation of protein
 To achieve this  must have the correct 1, 2, and 3 structure  if any of these
levels of protein structure breakdown start to have misfolding which can
contribute to dz states
 ALL proteins contain primary structure, majority proteins contain secondary &
tertiary structure, and some proteins contain quaternary structure
 Protein set to have quaternary structure if that protein consists of 2 or more
polypeptide chain
 Interaction of polypeptide chains w/ each other
 Simplest structure = dimer (2 subunits) = have 2 individual polypeptide chains 
can interact via non-covalent bonds & and sometimes covalent bonds (disulfide
bridges/bonds) that hold individual polypeptide together
 Whenever have quaternary structure those individual polypeptide chains = subunits
 Can be different or identical
 All proteins in our body can be put into two categories: Two types of proteins:
 1) Fibrous proteins
 Structural protein that consists of long fibers that play structural role in cell &
body
 Function: give cell/body structure
 Ex:
 Intermediate filaments = cytoskeleton
 Collagen = connective tissue  bone
 Keratin = hair & nails
 Alpha-keratin (dimer)
 Consists of 2 individual/long fiber – polypeptide chains – both
polypeptide chains are composed of right alpha-helixes = these
together intertwine to form the left handed coil = alpha coiled coil
 How are these 2 subunits held together?
 Covalent & non-covalent interactions
 van der Walls (London dispersion forces) b/w nonpolar side
chains of aa found on the 2 opposing subunits
 Ionic bonds  bonds b/w negatively & positively charged side
chains
 Hydrogen bonds
 Disulfide bond/bridge  covalent bond formed b/w 2 adjacent
cysteine aa’s  the more disulfide bonds have inside the
alpha-keratin – the stronger/more rigid that protein is

 2) Globular proteins
 Spherical shape
 Membrane bound/transfer proteins that allow movement of ions and
molecules across the cell membrane
 Ex:
 Hormones = insulin
 Enzymes = DNA polymerase (quaternary structure that contains many
subunits – globular protein)
 Hemoglobin = quaternary structure = tetramer = 4 polypeptide subunits (2
alpha & 2 beta) = O2 carrier inside blood
 Picks up O2 in lungs and moves O2 via circulatory system into cells &
tissues that need O2 to synthesize ATP
 Each subunit has prosthetic group – HEME = binds O2 via
oxidation/reduction rxn – heme group 1,2,3,4 – heme groups bind a
single O2 molecule each – can bind 4 O2 molecules per Hb
 Slight changes to quaternary structure of Hb can increase or decrease
affinity of Hb to O2
 Myoglobin = only contains tertiary structure because it consists of a single
polypeptide chain (not 4 polypeptide chains like Hb)

Structure Element Definition Subtypes Stabilizing Bonds

1°  Linear sequence of AAs in  NONE  Peptide (amide) bonds
chain
2°  Local structure determined by  α −helix  Hydrogen bonds b/w 
nearby AAs  NH & C=O
β− pleated sheets
3°  3D shape of protein  Hydrophobic  van der Waals forces
interactions  Hydrogen bonds
 Acid-base/salt  Ionic bonds
bridges  Covalent bonds
 Disulfide links
4°  Interaction b/w separate  NONE  van der Waals forces
subunits of a multisubunit  Hydrogen bonds
protein  Ionic bonds
 Covalent bonds

 PROTEIN CONFORMATIONAL STABILITY  Folding = chain  3D structure

 = ALL forces that help keep protein folded in right way  forces = 4 different levels of
protein structure 1-4 and solvation shell/environment that the protein is in
 Many proteins fold spontaneously  some need chaperone proteins
 Conformation = folded 3D structure – active form of protein
 Denatured (protein) = unfolded/inactive proteins
 Extreme heat
 [Salt]
 pH
 Protein only functional when in proper conformation (3D form)
 In addition to 4 levels of protein structure, another force stabilizes protein conformation =
solvation shell = layer of solvent that surrounds protein
 Protein w/ (+) charged exterior residues  protein in the watery environment of
interior of cell  solvation shell is layer of H2O right next to protein (H2O = polar
molecule)  have electronegative O atom w/ (-) charge leaving (+) charge next to H
atoms  electronegative O atoms are stabilizing all the positively charge aa residues
on exterior of protein
 How does protein become unfolded/inactive – denaturation?
 Temperature
 Heat = form of energy
 Add heat – breaks bonds
  When change the temperature destroy all the levels of protein structure
except primary structure
 Destroys….
 Secondary
 Tertiary
 Quaternary
 BUT the primary structure is still preserved
 pH
 add ACID – add a bunch of (+) charges – Ex: vinegar
 Break all of the ionic bonds that contribute to tertiary & quaternary structure
 When change the pH  have destruction of ionic bonds
 Ionic bonds  dependent on the interaction b/w (+) & (-) charges  when
add acid or base disrupt the balance b/w the (+) & (-) charges in the protein
 Chemical denaturant
 Disrupt hydrogen bonding w/in protein
 H-bonds contribute to secondary, tertiary, and quaternary structure – so these
will be disrupted if add chemical denaturant
 Add alcohol  all H bonds will be broken left w/ all linear polypeptide chains
 Enzymes
 Digestive tract  enzymes that break down proteins  take linear polypeptide
chain whose primary structure still intake  break bonds b/w individual aa –
break the peptide bonds  use as building blocks for own protein synthesis
 How enzymes can alter the proteins primary structure – and overall the proteins
conformational stability

 Anfinsen’s Experiments of Protein Folding

 In order to perform the 3D structure of polypeptide is stored in the specific sequence of
aa’s in that polypeptide  primary structure determines that conformation of that
protein
 Materials used in experiment
 1) 2 denaturing agents  molecules that breakdown the structure of protein  agents
used to breakdown the secondary & tertiary structure of protein
 Urea
 Used to break down the noncovalent bonds – Hydrogen bonds & Ionic bonds
– that hold together the secondary structure and part of the tertiary structure
 Beta-mercaptoethanol
 Used to break down the covalent bonds – disulfide bonds – that hold the
structure of tertiary structure together
 Uses oxidation/reduction rxn  reduces disulfide bonds and breaks down the
cysteine units into 2 individual cysteine amino acids
 What protein was used?
 Ribonuclease  enzyme/protein that catalyzes the breaking down of RNA molecules
in cells  have 124 aa’s in its primary sequence and in the tertiary structure it
contains 4 individual disulfide bonds
 Have 1 bond b/w 26 & 84th cysteine aa and have a 2nd bond b/w 40th & 95th have 3rd
bond b/w 65th & 72nd, and have final bond b/w 58th & 110th cysteine aa in ribonuclease
 3D structure/conformation describes the native structure/conformation of
ribonuclease
 Native structure/conformation = describes the biologically active structure of
enzyme
 Plan: Destroy the secondary & tertiary structure of ribonuclease by using these agents
and see under which conditions did the native structure of ribonuclease reform 
these 3 experiments proved that it is the primary structure (specific sequence of aa’s)
that gives the information to form that final 3D of polypeptide
 Experiment 1:
 Took active ribonuclease enzyme (native form) that contains secondary & tertiary
structure and mixed it w/ excess amount of Beta-mercaptoethanol and [8 molar]
of urea  the excess Beta-mercaptoethanol broke down all of those covalent
disulfide bonds & urea broke down the noncovalent bonds broke down that hold
the secondary & tertiary structure  @ end formed denatured enzyme – inactive
enzyme (not in active form)
 Removed 2 agents @ the same time  isolated denatured enzyme  saw that
because the denatured enzyme was in presence of O2, the O2 in air was able to re-
form the disulfide bonds – oxidized those disulfide bonds and because the urea
was removed @ the same time the noncovalent bonds also re-form and re-form to
the original secondary & tertiary structure of that enzyme
 Gives us evidence that it is the primary structure that dictates the proper formation
of tertiary structure of enzyme
 Experiment 2:
 Took that beaker that contained denatured enzyme as well as the excess Beta-
mercaptoethanol and urea and instead of removing these 2 agents @ the same
time, he 1st removed the Beta-mercaptoethanol and then after some time he
removed that urea
 Found that the enzyme that was formed was not in its biologically active state 
this enzyme was scrambled it contained the incorrect pairing of disulfide bonds
 This native form of enzyme contains these disulfide bonds – pairings – will
contain the improper linkages b/w cysteine molecules and that will create an
inactive molecule
 Why did this take place?
 It is the primary sequence – specific sequence of aa in that polypeptide that
dictates the type of noncovalent interactions that will exist on that peptide and
it is these noncovalent interactions that drive the correct information of
disulfide bonds
 If these noncovalent interactions cannot exist in that polypeptide, then that
polypeptide has no way of knowing what the proper disulfide bonds are that
have to be formed
 Because initial removed the beta before urea, saw that in denatured mixture
have the urea so the noncovalent bonds could not form or drive the correct
formation of those disulfide bonds, but because had urea in mixture, improper,
incorrect disulfide bonds were formed
  Initial removal of beta-mercaptoethanol created inactive enzyme was formed
because of the improper disulfide bonds were formed because those
noncovalent bonds could not dictate the formation of those proper disulfide
pairings
 Experiment 3:
 Saw if take scrambled enzyme and added beta-mercaptoethanol  because as
this catalyst  this catalyzed the breakage of these incorrect paired disulfide
bonds and because of thermodynamics form this native enzyme
 It is the conformation/structure of that native enzyme that is the most stable
thermodynamically  because of this we have breaking of this bonds and
formation of this native structure
 If take scrambled enzyme  beta break the incorrectly paired disulfide bonds
and this structure will form because it is the most thermodynamically structure

 Proteins & Protein Misfolding

 What happens if that polypeptide that begins w/ correct primary sequence folds
incorrectly?
 Final 3D structure of polypeptide is not correct
 3D structure of polypeptide that determines the function  function of final
misfolded protein will not be the same  these types of misfolded proteins are
biologically inactive  cells will either denature or refold them into correct
structure
 Structure = function
 Rare cases, misfolded proteins can  form prions  infectious agents

 Prion
 1) Aggregates of proteins that normal exist in body but have misfolded  insoluble
aggregates
 2) transmissible  1 organism to another
 3) insoluble  cells cannot break them down or denature them the same way it
normally does  cause death of body and then person
 Ex: PRP = normal protein found in brain  mutation PRP can misfold into PrPSC
 PRP consist of alpha helixes
 PrPSC has high content of beta-pleated sheets  because have high content these
molecules will have high potential of binding to other molecules that also contain
beta-pleated sheets
 Beta-pleated sheets consist of linear polymers of aa’s stacked on top of one
another  parallel to each other  have great potential to other beta-pleated
sheets via noncovalent bonds
 Can form aggregate (multi-unit of PrPSC) molecules when interact w/ each
other  form even larger fibers
 PrPSC infected misfolded proteins can transform normal proteins into
abnormal proteins  bind to multi-unit aggregate, transform them, and form
amyloid fibrous – amyloid fiber – form even large aggregates that cannot be
broken down by body’s cells  effect efficient and functions that take place
in cells (brain cells in this case)  proteins kills of nerve cells in brain and
 degenerates the mental capability and function of brain= Creutzfeldt-Jakob dx
(CJD)

 Alpha-Amino Acid Synthesis

 1) Gabriel synthesis
 Reason why start out w/ all other groups attached to key atoms  amid
prevented/protected from acting as nucleophile by having phthalimide group attached
to it & carboxylic acid protected w/ ethyl ester that is attached and alpha carbon
further activated by additional ester group @ top
 In presence of base and source of alkyl group “thad” will become alkylated
 Alkyl group substituted onto carbon aotm = alkylated step
 2) Hydrolyzed step - Acid hydrolysis
 Phthalimide group hydrolyzed along w/ 2 esters
 3) Add heat – decarboxylate
 Remove carboxyl group @ top
 2) Strecker synthesis
 More simple/elegant/efficient
 3 starting components
 1) Ammonia (NH3) = precursor for amino group
 2) potassium cyanide = precursor for carboxylic acid group
 3) Aldehyde or Ketone = serves as scaffold on which the amino & carboxylic
acid groups will be bound  provides the carbon that becomes the alpha carbon

ESTERIFYING ACID GROUP(s):

 Carboxylic acids can be esterified by rxn w/ alcohols in presence of Acid Catalyst
o Carboxylic acid in AA behaves same:

 Acid is a catalyst for esterification but remains acidic when rxn is over.
o Doesn’t “know” it is there ONLY as catalyst, so it will also react w/ alkaline
amino group  so needs to be shown as reactant.

Correct one

______________________________________________________________________________
1B. TRANSMISSION OF GENETIC INFORMATION FROM THE GENE TO PROTEIN

CENTRAL DOGMA: Kaplan: Sections 1.1 – 1.6 (Biochemistry Book) Sections 10.1 – 10.3
(Organic Chemistry Book)

DNA  RNA  PROTEIN

 Transcription = nucleus
 Translation = cytoplasm
 RIBOSOMES read off mRNAs  make proteins
 Proteins synthesized by RIBOSOMES

 EPIGENTICS
o Study of heritable changes in gene activity that are NOT caused by changes in
DNA sequence
o Same DNA sequence can be modified resulting in different phenotype without
changes to the underlying DNA sequence
o Allow transcription of certain genes within the genome depending on the cell type
o MECHANISMS: that produce these changes  explain why have same DNA in
each of cells of own body but those cells do not behave the same way
 DNA methylation
 Histone modification
 Ex: DNA in nucleus of muscle and skin cells is the same DNA  but
these two cells are different because the expression of that DNA is
modified by these epigenetic mechanism

 GENETICS (simple)
o Changes in phenotype are based on changes in genotype

 3 KEYS OF ENZYMES
 1) Biological molecule personality = structure + polarity
 2) E-S fit correlates w/ enzyme effectiveness & LOWER Km
 3) Inhibitors
 Competitive = BLOCK active site
 Noncompetitive = BIND allosteric site  conformational change of active site
 Uncompetitive = BIND ES complex  DECREASING [S]

 ENZYMES & CATALYSTS

 Carbonic Anhydrase = enzyme in blood & saliva
 How do enzymes make rxns go faster?
 Use catalytic strategies to push rxns along quickly
 1) Acid/Base Catalysis = when enzymes act as acids or bases  helps w/ proton transfer
 Acids & bases = proton donors & proton acceptors
 Keto-Enol Tautomerization
 Have proton moving from C atom to an O atom
 Since acids and bases are good proton carriers they can help this rxn go quickly
by moving that proton around instead of molecule doing it by itself
 2) Covalent Catalysis = when enzymes form covalent bond w/ another molecule, usually
their target molecule  covalent bonds = 2 molecules sharing electron  helps w/
electron transfer
 Decarboxylation rxn = carboxy or CO2 group is being taken off a molecule 
rxns have a lot of electrons moving around, so if have covalently bond enzyme
that could hold onto some electrons – be an electron carrier “sink” that would help
this rxn move faster
 3) Electrostatic Catalysis = deals w/ stabilizing charge
 DNA (-) charged polymer because of (-) charged phosphate groups in DNA
 If an enzyme had a metal cation on in (magnesium) could use it to stabilize the (-)
charge found in DNA and make it easier to work w/
 DNA polymerase (enzyme that allows DNA replication to occur)  needs to
found a way to counteract all of the (-) charge on DNA – magnesium ions used
here
 4) Proximity & Orientation Effect = increase frequency of successful collisions of
molecules we want reacting together = collisions happen more often
 In order for 2 molecules to react w/ each other, they need to physically collide @
some point
 Molecule A & molecule B will only react when they crash into each other
 Enzymes = able to bring 2 molecules close together, so these collisions happen more
often = making the 2 molecules happen more quickly
 Orientation of 2 colliding molecules in space is important  if molecule A & collide
but if 1 is not in correct position then the collision may not result in a successful rxns
 Enzymes also make sure that the 2 molecules will collide in the right orientation
 Increases the frequency of collision and helps to make sure that collisions are
successful & result in a rxn
 Summary
 1) Enzymes increase the rate of biochemical rxns – happen more quickly
 2) 4 catalytic strategies that enzymes can use
 1) Acid/Base Catalysis
 Helps w/ proton transfer
 2) Covalent Catalysis
 Helps w/ electron transfer
 3) Electrostatic Catalysis
 Deals w/ stabilizing charge
 4) Proximity & Orientation
 Increases frequency of successful collisions of molecules reacting together

 MCAT question of Day

 Carbonic Anhydrase catalyzes a rxn involving which molecules to form Carbonic
Acid?
 (A) H2O & CO2
  (B) H2 & CO2
 (C) H2O & CO
 (D) H2 & CO

 Answer: A

 purpose of rxn to help carry CO2 from tissues to lungs to get rid of  CO2 not
easily carried by Hb  want to CO2 into something that can be dissolved in
bloodstream – bicarbonate – dissolves easily in blood to easily transport CO2 to
lungs
 ** know rxn pathway
 CO2 + H2O - H2CO3 (carbonic acid)  HCO3- + H+
 Enzyme: carbonic anhydrase
 In blood – 85% of CO2 is carried as HCO3-
 Using Cl- shuttle – shuttle bicarbonate out of the RBC and into the bloodstream
where it floats to the lungs  in lungs this rxn goes in reverse  send CO2

 Enzyme = catalyze biologically rxns inside cells, w/out enzymes catalyzing the rxns cellular
processes would hold to rate that would make life impossible
 Fact 1: enzyme = biologically molecule that catalyzes/speeds up the rate of rxns
 Ex: Rb w/ carbonic anhydrase  allows conversion of CO2 into its polar form
bicarbonate ions  allows us to store the CO2 inside blood plasma  carbonic
anhydrase hydrates CO2 to produce carbonic acid that dissociates into H+ (polar ions)
& bicarbonate ions (polar ions)
 Carbonic anhydrase = efficient enzyme = increases the rate by 1 million compared to
uncatalyzed form = helps us transform nonpolar CO2 that cannot dissolve inside our
blood into a form that can be dissolved inside our blood  helps to effectively and
quickly get rid of CO2 in our lungs
 Fact 2: enzymes transforms 1 form of energy (we can’t use) into a more useful form
of energy (we can use)
 Ex: photosynthesis  plants have enzymes that transform and capture the energy
from light  transform energy stored in light into energy stored in form of glucose
(sugar)
 Fact 3: Enzymes do not act alone  they require cofactors = helper molecules that
help enzymes function effectively and efficiently
 Enzyme not bound to cofactor = apoenzyme
 Cofactor bound to apoenzyme = holoenzyme = enzyme bound to cofactor
 Cofactors in 2 categories
 1) Metal ions
 Zinc  for carbonic anhydrase
 2) Organic molecules – coenzymes = formed from vitamins
 Coenzymes = can bind onto proteins strongly or weakly
 Tightly bound coenzyme = prosthetic group
 Fact 4: Enzymes are efficient and specific
  Enzymes only bind to specific reactants/substrate and catalyze aa single rxn or a set
of rxns that are closely related to one another
 Enzymes are highly efficient and limit the # of unwanted products
 Carbonic anhydrase = ensures that we form only single product no unwanted
products
 Set of related rxns = trypsin = found in digestive system = digestive enzyme =
binds to polypeptides to proteins that we ingest into our body and carries out a set
of two closely related rxns  trypsin has single type of substrate and carries out
2 sets of rxns
 Rxn 1: Cleaves peptide bonds on the carboxyl side of lysine
 Rxn 2: Cleaves on carboxyl side of arginine amino acid
 Fact 5: Nearly all enzymes are proteins  some enzymes are also RNA molecules
 Certain RNA molecules also have ability to catalyze rxns
 Fact 6: Enzymes NOT used up in chemical rxns  if enzymes are changed/altered
in rxn, @ the end the enzyme will assume original shape and structure  enzymes
are not used up and remain unchanged @ the end of the reaction
 NOT enzymes during rxn are changed, their structure might be changed, but @ the
end of rxn when enzyme releases substrate it resumes its original structure and shape

 ENZYMES & ACTIVATION ENERGY

 Enzymes = use “catalytic strategies” = 4
 Molecules energy level = related to its stability
 Enzymes  reduce the needed activation energy so these reactions proceed @ rates that
are useful to the cell
 Lower energy state = more stable
 Higher energy state = less stable
 More unstable form it needs an input of energy to get there
 Transition state
 Highest energy point on path
 Most instability in entire rxn
 Different b/w energy level where start and the top of graph = delta G = free energy of
activation = amount of energy that A needs to have in order to break the rxn barrier to get
to point B
 Difference in energy b/w point A & B = standard free energy change (for entire rxn) =
represents the net change in energy levels between reactant and product = also the energy
that is released into environment once the rxn is over
 Rxns have products @ lower energy state than products = spontaneous rxn
 Free energy of activation energy – difference b/w point A and the transition state value -
that determines how quickly a rxn will go  usually delta G (standard free energy
change) is much higher than free energy for the rxn  which is why enzymes speed up
rxn by lowering the rxns activation energy
 Delta G double dagger = EA
 Standard free energy change = ERXN
 More stable w/ enzyme = rxn as whole will have lower activation energy  rxn has
transition state w/ much lower energy
 Enzyme does NOT change @ start or end
 
Starting & ending points are the same  enzymes are NOT used up when they
catalyze the rxn
 The only thing that changes is the path you take to get from A to B
 No permanent change to enzyme following the rxn
 SUMMARY
 1) Enzymes work by lowering the free energy of activation of a rxn  easier for
reactants to transition and form products  Free energy of rxn does NOT change
when use enzyme and when you don’t
 2) Despite change in pathway to get from A to B, the reactants and products DO
NOT change & enzymes DO NOT change equilibrium (of rxn)
 3) Enzymes are NOT consumed when they catalyze a rxn and the same enzyme can
catalyze rxns over and over again

 When we want to know how a certain rxn takes place, we want to study the thermodynamics
and kinetics of that rxn  Gibbs free energy – enthalpy and entropy – and activation energy
 Gibbs free energy = energy b/w reactants and products = where that equilibrium
will be achieved
 = describes how much energy can be released/used in that chemical rxn
 Reactants  products
 Products have lower free energy than reactants
 Take free energy of products – free energy reactants = Gibbs free energy = delta
G = how much energy is going to be released in this rxn  how much energy can
we use
 EXERgonic = (-) delta G = energy released = spontaneous rxn
 Ex: Combustion rxn
 Ex: breaking down ATP molecules  energy released to power body
processes
 Assume rxn has not reached equilibrium  can have (-) or (+) Gibbs free energy
 Reverse rxn
 Subject high free energy from low free energy = (+) delta G = ENDERgonic rxn
= nonspontaneous rxn = it will not take place unless a certain amount of energy
is inputted
 Ex: Synthesis of ATP molecules  to synthesis ATP have to input energy
 Gibbs free energy ONLY depends on the free energy value of the reactant and the
free energy of the product  if we know what the free energy of the products is and
the free energy of reactants, subject the 2 to find the Gibbs free energy
 Pathway from reactant  products does NOT change what the Gibbs free energy
is
 Does not matter if we take pathway 1,2, or 3 when go from reactants  products
 Gibbs free energy will NOT change
 If rxn that has enzyme and a rxn that has no enzyme  Gibbs free energy will be
exactly the same
 Catalyzed and uncatalyzed rxn will have the SAME Gibbs free energy value
 remains unchanged when the enzyme acts on that rxn
  enzymes when they act on chemical rxns, they do NOT affect the
Gibbs free energy value – they do not change the energy of the reactants
 nor do they change the energy of the products = difference delta G – Gibbs
free energy remains the same when enzyme is used or enzyme not used
 Enzyme does NOT change Gibbs free energy (∆G) – energy of reactants
and products, so difference – delta G – is the same
 What happens if Gibbs free energy = 0?
 Then no energy is being produced in that rxn that can be used in any useful way
 That rxn reached equilibrium  rate of forward rxn = rate of reverse rxn
 Neither spontaneous nor nonspontaneous

 Activation Energy (∆EA) = how quickly rxn takes place = how quickly that equilibrium
will actually be achieved
 Any rxn has some activation energy
 = amount of energy have to input/supplied for rxn to take place in forward or
reverse
 Energy barrier that must be overcome for rxn to proceed
 Enzymes LOWER EA by  stabilizing the transition state
 Reactants  products  activation energy = difference b/w energy of molecule found on
top of hill and the energy of reactant
 Enzymes stabilize the transition state  lower the energy of transition state 
decrease the activation energy speeds up chemical rxn
 Top most apex of hill = energy of transition state of chemical rxn
 Transition state = does not exist for long time because it has a very high energy
value  highest energy value in rxn  which is why the transition state does not
exist for long, so it is unstable – can’t isolate it because it quickly converts into the
products
 Describes how quickly a rxn takes place
 Spontaneous rxn = but it can take place slowly  if rxn takes place slowly = HIGH
activation energy
 Apex = energy of transition state
 Energy required to get to equilibrium (rate of forward & reverse rxns are the SAME)
correlates w/ ∆ G & is unchanged when enzyme present
 Activation energy is NOT the same thing as Gibbs free energy

 How does the enzyme affect the transition state?

  Lowers the energy of transition state  it makes this “hill/height” smaller and the
activation energy of rxn will be smaller
 If decrease the activation energy by stabilizing the transition state – speed up the rxn
– it is the activation energy – energy barrier – that determines the kinetics – speed &
rate – of chemical rxn
 Enzymes do NOT affect the equilibrium  no effect/do not change on Gibbs free
energy  do NOT increase or decrease how many products are forms @ end of rxn,
but they allow equilibrium to be achieved quicker by increasing the rate by
decreasing the activation energy

 Induced fit
 Catalyzed = smaller activation energy
 Energy of catalyzed rxn transition state = LOWER than energy of uncatalyzed rxn
transition state
 Most enzymes = proteins
 Substrates = any molecule will act on  reactants that enzyme will help turn into
products
 Active site
 Location on enzyme where substrates bind & where the rxn happens
 Enzymes have unique active site  bind to certain substrates
 Two pieces of clay that mold together
 Enzymes are specific to certain substrates – rxns
 1) Enzyme & substrate  have not come in contact yet
 2) Initial binding of enzyme to substrate  binding is not perfect  forces holding these
2 together are strong but not @ maximum strength yet
 3) Enzyme & substrate change shape some so they bind together tightly = induced fit =
rxn @ this point is @ full force
 4) Occurs after rxn completed  similar binding as stage 2  enzyme cutting substrate
in 2 parts
 5) Products of rxn released from enzyme  enzyme goes back to same state as stage 1

 Series of events – different steps in sequence of rxns

E & S separate
E & S bind = Enzyme-substrate complex = ES
Induced fit stage 3 above = transition state of rxn = point where enzyme most tightly
bound to substrate
 Substrate is not reactant or product = X = it is somewhere in b/w
 After rxn occurred – after transition state – E bound to 2 products P1 & P2
 E separated from 2 products P1 & P2
 Binding b/w enzyme & substrate  strongest @ transition states = induced fit =
enzyme & substrate molded together
 Sometimes enzymes bind more than 1 substrate  don’t always bind 1 substrate
 Ex: lactic acid fermentation
 Enzyme: lactate dehydrogenase  has space to bind 2 different substrates –
NADH & pyruvate
  Allosteric binding = any binding site outside of active site – bound by regulators -
activators or inhibitors
 Regulating molecule – inhibitor – may bind enzyme @ different location that the
enzymes activation site  changes shape of enzyme that affects enzyme’s ability to
catalyze the rxn  cannot bind intended substrate because they do not fit together
 Can bind regulators @ allosteric sites

 ENZYME CLASSIFICATION: 6 TYPES OF ENZYMES

MNEMONIC: Enzymes: Classification

Over The HILL LI’L HOT
Enzymes get rxn over the hill.

Oxidoreductases Ligase
Transferases Isomerase
Hydrolases Lyase
Isomerases Hydrolase
Ligases Oxidoreductase
Lyases Transferase

 1) Transferase
 Move functional group X from molecule A  molecule B
 “transfer” functional groups from 1 molecule to another
 Ex: Protein translation  aa’s bound to tRNA molecules transferred to
polypeptide chain
 Peptidyl transferase
 2) Ligase
 Catalyzes rxn b/w 2 molecule  A + B  AB (combine to form complex b/w 2)
 “ligate” or join 2 molecules together
 Ex: DNA replication
 DNA ligase
 3) Oxidoreductase = 2 type of rxn
 Transferring electrons from molecule A  B or from molecule B  A
 Move electrons b/w molecules
 Oxidase = Oxidizing  taking electrons AWAY from molecule
 Reductase = Reducing  giving electron TO molecule
 Together = Oxioreductases = catalyze BOTH forward & reverse rxns
 Ex: Lactic acid fermentation
 Electrons passed from NADH to pyruvate or lactic acid to NAD
 Lactate dehydrogenase (dehydrogenase = removal of hydride functional
group = removal of electrons) hydrides = hydrogen atoms w/ 2 electrons
 Isomerase
 Molecule converted to one of its isomers
 Ex: Glucose-6-P  Fructose-6-P
  Phosphoglucose Isomerase
 6) Hydrolase
 Use/react w/ H2O to cleave molecule into 2 other molecules
 Break bonds using H2O
 Ex: Hydrolysis rxn to peptide bonds
 Serine Hydrolase (protease)
 Serine key catalytic aa responsible for breaking peptide bonds
 Lyase
 Catalyze the dissociation of molecule w/out using H2O & w/out oxidation or
reduction
 Break bonds w/out H2O or oxidation
 Need to generate a double bond b/w 2 atoms or a ring structure in molecule in order
to work
 Ex: Cleavage of arginine succinate  arginine + succinate
 Argininosuccinate lyase
 Lyase catalyzes the breakdown

Enzyme type Function Example

Isomerase Catalyzes isomerization rxn Enzyme that converts cis
= INTRAmolecular double bond  trans
rearrangement of bonds double bond
Ligase Catalyzes joining 2 molecule Enzyme seals the gap b/w 2
adjacent Okazaki fragments
Transferase Catalyzes transfer of Kinase that adds phosphate
functional group from 1 group from ATP to protein
molecule to another substrate
Lyase Catalyzes breaking of Enzyme that breaks bond b/w
molecule w/out H2O 2 nucleotides w/out H20
Hydrolase Catalyzes breaking of Enzyme that breaks bond b/w
molecule by adding H2O 2 nucleotides by adding H20
Oxidoreductase Catalyzes transfer of Enzyme that transfers extra
electrons b/w molecule electrons from NADH
electron carrier to protein
substrate

 Cofactor/Coenzyme
 Bind to enzyme in order for enzyme to function properly
 Coenzyme = carrier molecule  transferring different things from 1 molecule to another
 Organic carrier molecules
 Organic = Primarily carbon based molecule
 Carrier = hold onto certain things for enzymes to make catalysis run smoothly
 Ex: NADH = electron carrier
  NADH  NAD+ + H- (hydride)
 NAD+ = can accept electrons causing molecule to become NADH  that can carry
electrons for enzyme
 Ex: Pyruvate + NADH  Lactic Acid + NAD+
 Lactate dehydrogenase using NADH as coenzyme to transfer electrons to pyruvate
 NADH = electron carrying coenzyme
 Ex: Co-enzyme A
 Acts as carrier molecule that holds onto acetyl groups
 Metabolic rxns  carry 2 carbon acetyl groups from 1 molecule another
 Cofactor = directly involved in the enzymes catalytic mechanism = don’t strictly carry
something = stabilize enzyme or substrate or help rxn convert substrates from 1 form to
another
 Ex: DNA polymerase = synthesis new DNA
 DNA (-) charged due to (-) phosphate groups around it
 DNA polymerase uses Mg2+ (magnesium ion) as cofactor  use its (+) charge to
stabilize (-) charge of DNA
 Instead of acting as carrier the magnesium ion cofactor is more directly involved
in catalysis and stabilizes DNA
 Vitamins = organic cofactors & coenzymes different cofactors & coenzymes
 Dietary cofactors & coenzymes
 Body cannot build them from scratch and need to get them from diet
 Vitamin B 3  Niacin  precursor for NAD+
 Vitamin B5  precursor for Co1
 Minerals = inorganic cofactors (inorganic = not carbon based)
 Magnesium (Mg2+)  DNA polymerase
 Calcium =  bones & teeth  can act as a cofactor but does not act as enzyme
cofactor here, it is important part of structure
 Not all minerals act only as cofactor

 Enzymes stabilize
 By speeding up chemical rxn  enzymes decrease the time that is needed for that
chemical rxn to reach equilibrium
 Enzymes = decrease time that is needed to reach equilibrium  but they DO NOT
change the equilibrium itself, they DO NOT change the energy of products or
reactants nor do they change the amount of products or reactants that is formed at
equilibrium
 (-) value = free energy released
 (-) Delta G = as long as have enough energy to overcome activation barrier, the
reactants will spontaneously  products are lower in energy and are more stable
 Enzymes DO NOT affect free energy value of products or reactants  delta G difference
b/w products and reactants will not be affected
 Because it is energy of products and reactants and difference of the energy b/w them
 determines the [ ]’s of product & reactants that will exist @ equilibrium because
the enzymes do not affect energy values they will not affect the [ ] of products and
reactants that exist at equilibrium
 Same [ ] of products and reactants will be formed in the presence or absence of enzyme
  If add enzyme the energy value of reactants and products does not change
 If the thermodynamics of products & reactants is not changed by enzyme, what is
changed?
 The kinetics of the chemical rxn is determined by the energy of the transition state
 Transition state = transient stage that exist b/w the reactants & products
 Reactants  products have to break bond b/w A and B and have to form bond b/w B
and C
 B moves away from A  electron density will move away and the bond will
break away (dashed line)
 B is approaching C  electron densities of 2 atoms is overlapping so begin to
form that bond (partially formed bond and partially broken bond) and because the
electron densities are not overlapping well  increase the energy of the transition
state  energy of transition state represents the highest possible free energy value
on the curve
 To calculate the free energy value of the transition stage of that molecule  take
the y coordinate value and subtract the energy of reactants  energy of transition
state – energy of reactants = free energy of activation = activation energy (barrier)
 When the enzyme takes these molecules  have location in enzyme = active site =
creates a microenvironment and binds to reactants  reactants move into active site of
enzyme  creating enzyme-substrate complex  enzyme stabilizes the partially broken
bond and partially formed bond = lowers the energy of activation  lower that free
energy of activation = increase rate that rxn takes place
 By binding substrates to active site enzyme stabilize the energy of transition state 
which stimulates breakage of old bonds and formation of new bonds to form that
product molecule
 The change in Gibbs free energy b/w products and reactants DOES NOT change
 Energy b/w transition state and reactants  is what changed
 Lowering in energy = means the difference b/w transition state & reactants is smaller w/
an enzyme  what makes the rxn go quicker  by adding enzyme decrease the time it
takes for equilibrium to establish  once reach equilibrium the same [ ] of products &
reactants are formed in the catalyzed and uncatalyzed state
 Maximum velocity of enzymes = maximum activity @ which the enzyme will operate
 If continue adding then all the active sites on all enzymes will be filled = entire activity of
the mixture of enzymes will be @ maximum velocity
 The curved line never will cross the max. enzyme velocity
 If have 100 enzymes only have 100 active sites = have excess of substrates and only 100
active sits have a maximal activity that only 100 active site
 As increase [substrate] enzyme activity increases up to certain value = maximum enzyme
velocity
 Represents = if we have 1000 enzymes each = 1000 active site = if have 1000 substrate
molecules all the active sites will be filled and that is the max activity of enzyme

 6 properties of active sites

 1) Active site = 3D region found on enzyme that is responsible for binding onto that
substrate
  Inside the active site consists of residues (aa that are responsible for binding onto
substrate) and also have catalytic groups – residues part of enzyme that actually
catalyze that rxn
 2) Active sites = responsible for stabilizing the transition state as well as forming the
bonds involved in that chemical/biological reaction  inside the active site have
residues responsible for stabilizing and lowering the energy of the transition state  that
speeds up the rxn and have catalytic groups that are responsible for stimulating the
breaking and forming of bonds
 3) Active sites create a microenvironment
 When it binds onto substrate it closes off and creates microenvironment that is
nonpolar  only time find H2O molecules in the active site is when the H2O
molecule is actually a participant (reactant) in that rxn  otherwise never find H2O
molecules inside the active site
 Active site environment = nonpolar
 Microenvironment brings the reactants close together and orients them in just the
right orientation for that specific reaction to take place  it also decreases the
likelihood that other reactions take place  decreases the likelihood that unwanted
products are formed
 Active sites created nonpolar microenvironments in which bonds can be formed
and broken very easily
 Helps prevent unwanted rxns
 4) Active sites only make up small portion of that overall enzymes, so even though
the enzymes is large, the active site is small compared to overall structure and size
 Residues in active site  found very far apart from one another on that primary
sequence of polypeptide chain  to bring those residues close together that enzyme
has to fold and form this 3D shape and to fold and bring those residues close together
have to fold many times to form the active site
 The entire enzyme creates a scaffolding system that supports and stabilizes that active
site that is used by enzyme
 Can have other sides on enzyme outside of active site that play a role in regulating the
functionality of the enzyme = allosteric sites
 Other portions of enzyme can also interact w/ different components in cell
 Ex: variety of proteins and enzymes in cell membrane that bind onto cell
membrane, so other sections of enzyme can be responsible for adhering and
binding onto cell machinery
 5) Active sites typically bind substrates reversible via non-covalent forces
 Binding = hydrogen bonds, hydrophobic interactions, and van der Wall forces = non-
covalent forces = ALL promote the reversible binding b/w active site and substrate
 Reversible binding = once substrate binds to active site and once convert that
substrate into product it can release itself and move away from active site it will not
remain bound to active forever
 6) Active sites have structures complementary to corresponding substrate  For
non-covalent interactions b/w residues to be strong enough, the distance b/w bonds –
molecules forming these bonds – has to be short enough
  In order for substrate to actual get close to active site to get close enough to form non-
covalent bonds, the shape of that substrate has to be complementary to shape of
enzyme
 Substrate must fit into active site of enzyme

 Lock & key

 Even before they actually bind, what this model tells us, the substrate moves into active
site and they form those non-covalent interactions
 Induced fit
 Active site of enzyme is not exactly complementary to substrate  when binding takes
place, the enzyme conforms to the structure of that substrate
 The enzymes active site induces the shape and once the binding takes place the active site
conforms and takes the complementary shape of the substrate
 Shape of the enzymes active site is not exactly complementary but upon binding of
substrate to active site  binding causes active site to become complementary to that
substrate
 When the binding takes place, the active site of enzyme and substrate itself change shape
and they both conform into shape into which each one is complementary to the other
 Substrate becomes complementary to active site
 Active site become complementary to substrate

 Enzymes work best in specific environments  pH & Temperature

 Alpha-amylase
 Pepsin
 Effects of changing pH of enzymes environment
 DNA polymerase
 Under normal pH – hold onto Mg ion through electrostatic interaction b/w Mg2+ and
Asp residues  that is deprotonated and negatively charged at normal pH
 Reduced pH = Asp residue = protonated due to pH dropping
 Protonated form = Asp no longer has negative charge and cannot hold onto Mg2+ and
DNA polymerase cannot do its job right in low environment
 Keeping enzyme @ appropriate pH is essential to its function
 Proteins need to fold into secondary, tertiary, and quaternary structures in order to be in
their functional form
 Significant changes to protein temperature can disrupt proteins folded geometry and
cause it to lose its functionality
 1) Enzymes function only under very specific environmental conditions  different
enzymes function ideally in different environments
 2) Changes to enzymes environment – pH & temperature  can lead to a loss of
enzyme functionality

 Non-enzymatic protein function

 Unique characteristic = can bind various biomolecules & bind specifically & tightly
 How they can form vast array of functions
 2 classes of proteins = can have characteristics of both
 1) Enzymatic proteins = enzymes
  Catalyze chemical rxns that help sustain life
 Help build up and break down things
 Help accelerate the rate & specificity of rxns
 Ex: DNA polymerase or enzymes in saliva = amylase  starch into sugars

 2) Non-enzymatic proteins = non-enzymes

 Proteins that carry out functions that require capacity to bind, but not to catalyze
reactions
 Ex: receptors/ ion channels / transport proteins / motor proteins / integral part immune
system = Antibodies
 Receptors/Ion channels
 Receptors = proteins that receive or bind signaling molecule
 Receptor binds signaling molecule – ligand – induces chemical response in cell
 Ex: Insulin receptor & insulin
 Ligand = insulin that binds to insulin receptor that leads to cascade
 Ion channel = expands the lipid bilayer
 Certain ions can enter or exit cell
 Transport proteins = bind small molecules & transport them to other locations
 Have to have high affinity for their ligand when ligand is present in high
concentration
 High concentration ligand = high affinity of protein for ligand
 Low concentration ligand = low affinity of protein for ligand
 Ex: Hemoglobin
 Picks up O2 in lungs – high in O2 – then delivers O2 to tissues where there is low
concentration of O2
 Motor proteins = cellular motility
 1) Myosin = generating forces exerted by contracting muscles
 2) Kinesin = intracellular transport
 3) Dynein = intracellular transport and motility of cilia (extensions of cell that
extend out)
 Antibodies (Ab)
 Protein components of adaptive immune system
 Antigen from foreign substance is the antibody’s ligand
 Antibody’s affinity for its target antigen is high/strong

 Covalent modifications to enzymes  a way to regulate & control activities of enzymes &
change functionality of proteins
 Not all enzymes are proteins
 Transfer a functional group/moiety from one molecule to that target enzyme/protein 
covalently attach a group onto that enzyme  changes activity of that enzyme or
functionality of protein  it can either turn on or turn off the activity of that enzyme
 Many types of covalent modifications that take place inside cells  modify proteins in
many ways
 Proteins
 Aa polymers w/ primary, secondary, tertiary, and quaternary structures
 Non-proteins
  Inorganic metals
 Mg2+
 Small organic molecules
 Flavin
 Covalent modifications = forming or breaking covalent bonds
 Translation = synthesis of aa polymer
 Post-translation = events that take place after initial synthesis
 Small = modifications that involve small functional groups being added or removed from
an enzyme
 1) Small post-translational modifications
 1) Methylation
 Addition of methyl group – CH3 to protein
 2) Acetylation
 Addition of acetyl group
 Ex: Acetylation of Lysine
 Extra aa on side chain that can act as base and carry positive charge
 Acetylate lysine group – add acetyl group to amino nitrogen = covalent
modification  electron withdrawing effect of the acetyl group will prevent
that nitrogen from carrying a positive charge and modify the behavior of that
amino acid  loss of positive charge can change properties to amino acid 
change the lysine electrostatic interactions with other charged molecules and
lysine’s acidity and basicity
 3) Glycosylation
 Addition of sugar molecules
 2) Zymogens = require covalent modification to become active  inactive proteins that
require covalent modifications to become active
 Ex: pancreas releasing trypsinogen (zymogen) -ogen = inactive form of protease
enzyme and once in intestine it is covalently modified that converts it to its active
form trypsin by enterokinase  only allows it to break down proteins in the
intestine
 Suicide inhibition = enzyme inhibitors that permanently bind their target
 Covalently bind the enzyme and prevent it from catalyzing rxns  form covalently
linkages to proteins they rarely unbind  once they bind that is it for them

 PROTEIN MODIFICATIONS
 After polypeptide chain formed  3D shape = can call it protein  but still need to make
protein modifications

 2 TYPES OF PROTEIN MODIFICATIONS

 1) Co-translational modifications
 Changes that happen to polypeptide while it is being translated
 Ex: Acetylation
 1st aa in polypeptide (usually methionine) is going to be removed  in its
place an acetyl group is put in
 Happens to majority of eukaryotic proteins
 2) Post-translational Modification
 Most protein modifications fall into this category
 Modifications happen after translation
 Location: ER & Golgi
 Ex: Glycosylation – structure of enzyme
 Adding of carbohydrate to a protein
 Happens to proteins that end up being embedded in cell membrane
 Helps to identify different types of cells
 Use this in ABO blood groups
 4 RBCs and each belong to different person > RBCs have proteins
embedded in their surface w/ different carbohydrate groups attached to
them
 Blood type A w/ that specific carbohydrate attached to it
 Blood type B w/ different specific carbohydrate attached to it
 Blood type AB w/ the specific carbohydrate for A and B attached to it
 Blood type O w/ does not have any carbohydrates of either category
attached to it
 How glycosylation is used in identification of different types of cells
 Ex: Lipidation – structure of enzyme
 Adding a lipid to a protein  protein attached/embedded to cell membrane
 Ex: GPI anchor  lipids that help to attach or tether proteins to cell
membrane  GPI plunges into hydrophobic region so lipid which is
hydrophobic attaches well to inner part of cell membrane that is also
hydrophobic
 Structure cell membrane  hydrophilic – polar heads on outside and
hydrophobic – nonpolar tails inside
 Location: ER & Golgi
 Ex: Phosphorylation - activity & function of enzyme & dephosphorylation 
Covalent modification  effective & efficient
 Process regulated by enzymes  protein kinases  enzyme responsible for
regulating phosphorylation
 Protein kinases  catalyze the transfer of terminal phosphoryl group from
ATP onto hydroxyl containing group (side chain) found on that target protein
 3 residues capable of using the hydroxyl group to accept that phosphoryl
group 1) serine 2) threonine 3) tyrosine
 Because they all contain hydroxyl groups on side chains
 What is the source of this phosphoryl group? We have a high abundance of
high energy ATP  contain the phosphoryl groups  source of the
phosphoryl
 Locations: inside the cell where the [ATP] is high  cells outside the cells do
not undergo phosphorylation because outside the cells ATP is not abundant
 Why is phosphorylation so common? More common methods of covalent
modifications
 1) Phosphorylation process gives a Net (-) Charge
  In the process of phosphorylation transform a neutral residue to a
negative charge modified residue
 Because of the presence of the negative charge it can break the old
interactions and form better/stabilized interactions
 If this negative charge residue is found inside the active site of the
enzyme, the presence of this modified phosphoryl group it forms new
interactions w/ the substrate molecule and can change the rate the
enzyme catalyzes that rxn
 Net (-) charge is what allows these new electric interactions to actually
take place b/w this residue and that target substrate molecule
 2) As result of modified (-) charge found on this residue  because of
this (-) charge it gives this modified residue a high potential to form
hydrogen bonds  the negatively charge oxygen atoms of
phosphorylated side chain residue can form hydrogen bonds w/ target
enzyme or target substrate molecule
 This can increase the specificity of the interaction b/w the active site of
enzyme and that substrate molecule
 3) Protein kinases are able to easily adjust the rates/kinetics of this rxn 
speed up rxn or slow down rxn
 rxn can take place quickly or over a long period of time
 rate @ which it takes place & rate @ which protein kinase catalyzes
rxn depends on the conditions found inside our cells
 4) Inside cells have high energy ATP that act as source of that phosphoryl
group
 Phosphorylation requires the transfer of phosphoryl group onto
residue of that enzyme  ATP molecules used to transfer that
phosphoryl molecule
 5) Amplification takes place whenever protein kinases are present
 Single protein kinase enzyme can catalyze many enzymes @ once
 Protein kinase amplifies all these different type of rxns & that speeds
up the # of substrate molecules that are made into products
 Activated protein kinases can be used to regulate many different
enzymes and many different pathways  leads to amplification of
final product
 Protein kinases  single one usually catalyzes many individual
enzymes and catalyzes many rxns @ once = amplifies effects and amt.
of product we form @ end
 6) Releases large amt. of free energy
 Dephosphorylating ATP molecule  breakdown of ATP molecule =
exergonic  releases energy
 This process is thermodynamically stable  because the products are
so much more stable than reactants this rxn takes place in single
direction and reverse rxn does not take place (not @ high rate)
 Energy of product is lower than energy of reactants
 Dephosphorylation of ATP = exergonic rxn  release a lot of free
energy
  Free energy that is released in the dephosphorylation of ATP is stored
inside phosphoryl group protein complex
 Products are more thermodynamically stable than reactants  shift rxn
all the way to the product side
 Products lower in energy than reactants
 7) Phosphorylation is reversible
 If phosphorylation turns on proteins activities have to be able to turn
off activity of enzyme by removing that phosphoryl group
 Reverse this rxn use different enzyme = protein phosphatases = used
to reverse the effects of protein kinases and remove the phosphoryl
group = dephosphorylate
 Can activate or deactivate the activity of the enzyme
 Protein kinase = phosphorylation
 Protein phosphatase = dephosphorylation
 Adding of phosphate group to protein or enzyme = phosphorylation
 Removing of phosphate group from protein or enzyme = dephosphorylation
 through the hydrolysis rxn  use H2O to remove the phosphate group and
reform the enzyme that contains the original residue
 Ex: Sodium-Potassium Pump = enzyme = Na+/K+ ATPase  this
enzyme/protein is responsible for maintaining the proper osmolarity of
Na+ and K+ ions in and out of cell
 How phosphorylation regulates this protein?
 1) 3 receptor site on enzyme for Na+ ions & 2 receptor site on enzyme
for K+ ions  Na+ ions attach to receptor sites on enzyme
(intracellular space – cytoplasm)  Na+ ions coming from cytoplasm
 when the receptor sites are full it causes an ATP molecule to break
down into ADP and this phosphate will attach itself to the protein =
phosphorylation  when this protein gets phosphorylated –
phosphate group gets attached to it - it causes conformational change
and protein now faces the outside of the cell  protein released Na+
ions into the extracellular space
 2) Still phosphorylated protein  K+ ions in outside of cell attach
themselves to protein on the receptor sites  when these receptor sites
are full it causes a different change  phosphate group is removed =
dephosphorylation  phosphate group ends up inside the cell 
when protein is removed causes conformational change of protein and
now it faces the inside of the cell  K+ ions are released into the
inside part of cell
 Phosphorylation & dephosphorylation regulates the activity of the cell
 Want HIGH [Na+] & LOW [K+]  outside cell
 Want LOW [Na+] & HIGH [K+]  inside cell
 For every 3 Na+ pumped out  2 K+ pumped in  how these
concentrations are maintained
 Ex: Methylation - activity & function of enzyme
 Ex: methylation of histones
  Histones = proteins that DNA wraps itself around  found in
chromosomes and help package DNA
 Methylating & demethylating histones  helps turn certain genes on &
off
 Ex: Proteolysis – activity & function of enzyme
 Take protein and activate it need to cut it (ex: insulin needs to be cut twice
before it is activated)
 Way to activate zymogen is by cutting it = proteolysis
 Way many proteins are activated
 Ex: Ubiquitination
 Add ubiquitin to another protein
 Marks protein for degradation  destroy protein and recycle different parts
______________________________________________________________________________

GLYCOLYSIS:

= breakdown of glucose (6-carbon sugar) to pyruvate (3-carbon sugar)

 NET YIELD = C7  2 ATP + 2 NADH

 HEXOKINASE
o = ENZYME that – c6 traps – glucose in cells by converting it to c6 glucose-6-
phosphate via addition of a phosphate from c6 ATP
o INHIBITED by: c6 product glucose-6-phosphate
 GLUCOKINASE
o = c6 liver form
o INDUCED by: c6 insulin
 PHOSPHOFRUCTIKINASE-1 = PFK-1
o c1 rate-limiting enzyme = of GLYCOLYSIS
o c1 fructose-6-phosphate = phosphorylated to c1 fructose 1,6 biphosphate
using c1 1 ATP
o INHIBITED by c1 ATP & c1 citrate
o ACTIVATED by c1 AMP
o ONLY PFK-1 = CONSUMES ATP in FORWARD rxn of glycolysis
o In LIVER:
 c1 insulin = STIMULATES
 c1 glucagon = INHIBITS  indirectly via c1 PFK-2 produce c1 2,6-BP

 GLYCERALDEHYDE-3-PHOSPHATE DEHYDROGENASE
o = oxidizes c2 glyceraldehyde 3-phosphate to the HIGH energy intermediate c2
1,3-bisphosphoglycerate by REDUCING c2 NAD+ to c2 NADH and adding one
c2 free phosphate (Pi)
o Needed for c2 FERMENTATION to continue

 3-PHOSPHOGLYCERATE KINASE
 o = transfers the phosphate from c3 1.3-bisphosphglycerate to ADP  forming
c3 ATP & c3 3-phosphoglycerate  this is the c3 FIRST substrate level
phosphorylation of glycolysis

 PYRUVATE KINASE
o = transfers phosphate from c4 phosphoenolpyruvate (PEP) TO ADP forming
c4 ATP and c4 pyruvate  this is the c4 SECOND substrate level
phosphorylation of glycolysis
o ACTIVATED by c4 fructose 1,6-bisphosphate from PFK-1 rxn via c4 feed-
forward mechanism
o PRODUCE ATP

 LACTATE DEHYDROGENASE
o = rate limiting enzyme c5 fermentation
o REDUCES c5 pyruvate TO c5 lactate and oxidizing c5 NADH TO c5 NAD+
replenishing the c5 NAD+ for glyceraldehyde-3-phosphate dehydrogenase
 DIHYDROXYACETONE PHOSPHATE (DHAP)
o = Glycolysis intermediate of glycolysis used in synthesis of triglycerides
formed form either F-1,6BP OR G3P

** What are the 3 irreversible enzymes of glycolysis?

1) c8:: Glucokinase OR Hexokinase
2) c8:: PFK-1
3) c8:: Pyruvate Kinase
TCA Cycle
** Know structures of TCA  including which carbons go where
** Know glucose, pyruvate, and acetyl-coA structures & # of carbons
INSULIN
 Peptide hormone = secreted by
pancreatic beta cells
 PROMOTES anabolic (synthetic
pathways)  energy requiring
 Binds receptor tyrosine kinases:
o Liver
o Muscle
o Adipose tissue

PANCREAS
 Islets of Langerhans
o Beta cell: secretes insulin
o Alpha cell: secretes glucagon
(opposes insulin)
GLUCAGON
 Peptide hormone = secreted by alpha cells
 Counterregulatory hormone = opposes:
o Insulin
o Epinephrine
o Norepinephrine
o Cortisol
o Growth hormone
 Counterregulatory hormone = promotes:
o Catabolic pathways in body that oppose actions of insulin
 Binds G-protein coupled receptors (GPCR) in:
o specific tissues – LIVER
o NOT in muscle
 SHORT ½ life
 SINGLE polypeptide chain
 Preproglucagon  Glucagon
o cleaved to produce diff. products in deff. tissues
 intestinal GLP-1
 Active secretion:
o Primary stimulus
 LOW blood glucose
o Stress hormones  override alpha cell’s response during physiologic stress
 Norepinephrine
 Epinephrine
o AAs
 HIGH protein meal stimulates glucagon secretion  counters insulin
secretion

INSULIN vs. GLUCAGON

 Glucose levels maintained in tight range
 Healthy adult fasting blood glucose:
 GLUCAGON MECHANISM OF ACTION
o Hepatocytes = receptors for glucagon & epinephrine
 Counterregulatory hormones control same processes, but response to other stimuli 
Ex: Stress
 Muscle cells = ONLY receptors for epinephrine NOT glucagon

1) Binds G-protein coupled receptor on hepatic cell

2) Alpha subunit exchanges GDP FOR GTP
3) Alpha subunit ACTIVATES adenylyl cyclase  which ACTIVATES PKA
4) PKA phosphorylates downstream enzymes
5) ACTIVATES glycogen & lipid breakdown & gluconeogenesis
6) INHIBITS:
 Glycogen
 Protein
 Lipid synthesis

 Long-term response:
o INCREASES transcription of
gluconeogenic enzymes
 Clinical Correlation
o Hypoglycemia
INSULIN QUESTIONS

1) Which of the following is the correct pathway for insulin secretion?

A. High blood glucose  glucose enters Beta cells  increase in ATP  ATP inhibit K+
channels  membrane depolarizes and Ca++ channels open  influx of Ca++ triggers
insulin secretion
B. High blood glucose  glucose enters Beta cells  increase in ATP  ATP opens K+
channels  membrane hyperpolarizes and Ca++ channels open  influx of Ca++ triggers
insulin secretin
C. High blood glucose  glucose enters alpha cells  increase in ATP  ATP inhibit Ca++
channels  membrane hyperpolarizes and K+ channels open  efflux of K+ triggers
insulin secretion
D. High blood glucose  glucose enters alpha cells  increase in ATP  ATP inhibit K+
channels  membrane depolarized and Ca++ channels open  influx of Ca++ triggers
insulin secretion

ANSWER
 A
 Blood glucose enters beta cells through GLUT2  glucose is converted to G6P by
glucokinase & goes through cellular respiratory, resulting in an INCREASE in
intracellular ATP  ATP INHIBITS ATP-sensitive K+ channels  results in
DEPOLARIZATION of cellular membrane  OPENS Ca++ channels  INTRACELLULAR
Ca++ INCREASES  promotes insulin secretion

2) Which of the following statements about insulin is incorrect?

A. It is secreted by pancreatic beta cells.
B. It upregulates catabolic pathways
C. It binds a tyrosine kinase receptors
D. All of the above are correct

ANSWER
 B
 INSULIN = UPREGULATES anabolic pathways  pathways that require energy

3) Which of the following portions of proinsulin is not secreted?

A. A-chain
B. B-chain
C. C-peptide
D. All of the above are secreted

ANSWER
 D
 Proinsulin comprises:
o A- chain
 o B-chain
o C-peptide
 In Golgi apparatus  C-peptide is cleaved off  produces ACTIVE insulin
comprising the A & B chains
 C-peptide still secreted along w/ ACTIVE insulin
o C-peptide = marker of insulin production & secretion  due to longer ½ life

4) Which of the following statements is correct?

A. GLUT4 can be found primarily in beta cells and hepatocytes
B. Glucokinase catalyzes the conversion of glucose to glucose-6-phosphate in
muscle tissues
C. Insulin has a half-life of hours
D. The high Km of glucokinase and GLUT2 contribute to their ability to serve as glucose
sensors.

ANSWER
 D
 GLUT4 = primarily found in adipose & muscle tissues
 GLUCOKINASE = found in beta cell & hepatocytes

 ENZYMES = Biological Catalysts w/ properties:

o LOWERS activation energy of rxn
o Affects the kinetics of rxn, but NOT the thermodynamics ∆ G OR equilibrium
constant
o Regenerates itself during catalytic cycles  regeneration = REDUCES the amt.
of protein that a cell has to make in order to carry out biochemical rxns

o Rxn rate = rate @ which reactants are consumed OR rate @ which products are
made

 ENZYME STRUCTURE & FUNCTION

o Structure determines function
  Change in structure => change in function

 REVERSIBLE RXNS
o A B  molecule undergoing rxn in BOTH direction @ rate of k1 =
AB and k2 BA

 k1 > k2  rxn will favor AB direction  end result = LARGER [ ] of B than
A @ equilibrium

 EQUILIBRIUM
o = proportions of A and B such that [ ]’s of A and B remain the same
 o substrate can turn into the product and the product can turn back into the
substrate  Le Chatelier’s principle
o When a rxn is @ equilibrium that means that it has reached the balance of
substrate and product [ ][s that makes it “happy”
o NET rxn does NOT proceed in either direction
o ** @ equilibrium  forward & reverse rxns are still continuing, but @
the same rate, so there is NO NET change in product vs. substrate [ ] 
NET rxn produces NO change in the proportion of A vs. B
 Enzyme catalysis
o A & B have a certain amount of energy, and in order to get from A to B
have to go through a transition state  which requires a lot of energy to
reach
o Enzymes do NOT affect the equilibrium of a rxn  adding an enzyme
will NOT shift the equilibrium [ ] to suddenly favor more A than B if the
natural equilibrium of the rxn is to favor more B than A

ENZYME KINETICS:
MNEMONIC:
Enzyme kinetics: competitive vs. non-competitive inhibition

With Kompetitive inhibition: Km increases; NO change in Vmax

With Non-kompetitive inhibition: No change in Km; Vmax decreases

** IS AN ARGININE OR A VALINE MORE LIKELY TO BE CATALYTICALLY

IMPORTANT AMINO ACID BASED ON ITS STRUCTURE AND CHEMICAL
PROPERTIES?
 ARGININE
 Valine does NOT have any atoms that are highly electrophilic or that want
to serve as nucleophiles

 Enzymes ARE NOT used up when they catalyze rxns

 AB
o simplify rxn
o Rate of change = k[A]
o k = rate constant dependent on the environment
 E + S  ES  E + P = Enzyme-substrate binding
 E = enzyme
 S = substrate
 P = product
o 2 rxns going on  each have their own rate equation
o Rate 1 = K1[E][S]
 has 2 reactants
o Rate 2 = K2[ES]
  has 1 reactant
o Rate of rxn = speed the rxn goes @ = V = speed  for entire rxn SP
d [P]
 V=
dt
o Increase the RATE to get new product  do this by:
 Increase [S]
 Increase [enzyme]
o Assume that K value = constant & CANNOT be changed
 Assume that total [enzyme] = constant
o Ex: Only have 4 enzymes that work @ a speed of 10 rxns/sec  absolute
maximum rate of rxn = 40 rxns/sec = Vmax
 Even if INCREASE [S] still a Vmax only so much we can increase rate of rxn by
increasing [S]
 As [S] Increase the rate levels off as approach Vmax value

 k = rate constant
 V = Rate = speed rxn proceeds
 Enzyme catalysis is divided into:
o 1st rate = enzyme BINDING to substrate
o 2nd rate = substrate FORMING product + enzyme (since enzymes are NOT
consumed in rxn)

1) Solutions are behaving ideally: 2 steps

 1st STEP: Binding of S to E
 2nd STEP: Transitions from S  P that the enzymes help = formation of
product
 Each step has a distinct rate
DO NOT have any external factors messing things up
2) 2 big constants STAY constant
 [E] is NOT changing  from protein synthesis and degradation
 Rate constant = k  is NOT changing from environmental factors  like temperature

3) S  P does NOT for product w/out help of enzyme  Negligible

 k1 = ES complex forms
 k2 = ES complex either dissociate OR turn into E + P @ k3
 k3 = turn into E + P

LINEWEAVER-BURK

 X-axis = 1/[S]
o = changes from [S] on Michaelis-Menten plot
 Y-axis = 1/V0
o = changes from V0 on Michaelis-Menten plot

 MICHAELIS-MENTEN
o V1 = Vmax [S]
-----------------
 Km + [S]

o V = rxn rate  amt. of product formed per unit time  depends on:
 2 primary LOCAL environment conditions that affect enzyme activity:
Rxn conditions (in body):
 1) pH
o AAs often found @ active site -charged AAs-
  these AAs have specific pKa for their sidechain
functional groups  depending on pH of the
solution, these functional groups can be protonated
OR deprotonated
 PROTONATION status important = determine
whether OR not enzyme can carry out catalytic
function
o Enzyme active site residues MUST have proper
protonation status
 2) Temperature
o INCREASING temp.  INCREASED kinetic energy of
enzymes & substrates  enzyme MORE likely to
encounter one another in correct orientation
 aka: molecules are bouncing around w/ greater
speed
o Temp. gets TOO HIGH = denatures enzyme  CANNOT
catalyze rxn
o ** Enzyme-catalyzed rxns = velocity doubles every 10℃
increase in temp. until optimum temp. reached
o [E] = held CONSTANT
o [S] = INCREASED
o V0 = initial velocity (rxn rate)
o Vmax = maximum rxn rate  speed of ALL the enzymes combined
o [S] = concentration of substrate (controlled variable)
o Km = amt. of substrate needed for enzyme to obtain ½ of its maximum rate of rxn
(Michaelis-Menten constant)
 = Binding affinity of the substrate for enzyme
 Km = intrinsic property of enzyme-substrate system & CANNOT be
altered by changing the [S] OR [E]
 LOW Km = HIGH affinity for substrate
 HIGH Km = LOW affinity for substrate
o Ka = association constant of the enzyme-substrate complex
 LOW Ka = LOW affinity for substrate (enzyme-substrate complex = LESS
stable)
 HIGH Ka = HIGH affinity for substrate (enzyme-substrate complex =
MORE stable)
o Kd = dissociation constant of the enzyme-substrate complex
  LOW Kd = HIGH affinity for substrate (enzyme-substrate complex =
MORE stable)
 HIGH Kd = LOW affinity for substrate (enzyme-substrate complex =
LESS stable)
o kcat = # of substrate molecules each enzyme converts to product per unit time =
turnover #
o Ki = binding affinity of the inhibitor
 Ki > 1 = inhibitor has HIGHER affinity for enzyme than substrate
 Ki < 1 = inhibitor has LOWER affinity for enzyme than substrate
o IC50 = ½ max. inhibitory concentration
  Tells how much of a drug is needed to inhibitor a biological process by
50%

 ** LOWER [E] & HIGHER Vmax  each enzyme is individually working it’s ass off
 ** Michaelis-Menten needs “saturating conditions”  so substrate >> enzyme

QUESTION
1) Which experimental condition is NOT necessary to achieve reliable data for Michaelis-
Menten enzyme kinetics?
A. Initial velocity is measured under steady state conditions.
B. Solution pH remains constant at all substrate concentrations.
C. The concentration of enzyme is lower than that of substrate.
D. The reaction is allowed to reach equilibrium before measurements are taken.

ANSWER
 D
 To measure Vmax – when graph plateau – [S] >> [E]

Michaelis-Menten 3 assumptions: ** ALL of these are true @ start BUT may NOT be @
end

1) Free Ligand Approximation: [S] >> [E]

 If you have equal amts. of S & E OR less S  formation of ES complex would deplete
[S]  slowing down the rxn rate = Michaelis-Menten equations/graph NO longer valid
 [S] = constant  this approximation ONLY true during initial rxn phase  be4
significant amt. of S converted to P
 S can be depleted when binds the enzyme to form ES complex  to ensure ES
formation does NOT impact [S]  total [E] in solution NEEDS to be smaller than any
[S]
2) Steady State Assumption: Conversion of ES complex to product = constant rate
 If the amt. of substrate runs low  then rate would slow down (rate equation NO longer
valid)
 STEADY STATE = [ES] complex = constant  meaning formation of ES = loss of
ES
o ES formation = ES loss
  [ES] = constant over rxn course  allowing rate of product formation to remain
constant
 Once depleted  ES levels DECREASE = RXN SLOWS
 Rate of ES formation = ES breakdown (to E & P)
 NOT necessary for ES complex to form products  REVERSE rxn allows them to
dissociate back into individual enzymes & substrates
3) Irreversibility Assumption: Rxn ONLY occurring FORWARD direction: E + S  ES 
E+P
 True @ beginning of rxn BUT may NOT be true if the rxn has been going on for a while
& S levels run low (LeChateliers principle)
 Rxn proceeds = FORWARD DIRECTION ONLY
o  P NOT converted back to S
 Once enough P accumulates  reverse rxn occurs @ non-negligible levels  SLOWS
NET RATE of P formation

4) ONLY initial rxn velocities used in the analysis of enzyme rxns

 Rate of rxn @ zero time - @ that time [P] is small  rate of back rxn from P to S
= ignored

RELATIONSHIP B/W VELOCITY TO [E]

 Rxn rate = directly proportional to [E] @ ALL [S]
o  if the [E] halved = V0 REDUCED to ½
 ORDER OF RXN:
o When [S] << Km  V = roughly proportional to [S] = 1st order rxn
rate
o When [S] >> Km  V = constant & equal Vmax  rxn rate =
independent on [S] = 0 order rxn rate

 FEEDBACK REGULATION
o Positive Feedback = promotes/enhance rxns
o Negative Feedback = inhibits/reduces rxns

 COOPERATIVITY (aka + OR – cooperative binding) = SIGMOIDAL CURVE

o Binding of substrate to one subunit allosterically INCREASES the affinity of
other subunits for substrate = cooperative case  occurs when enzyme
subunits are STRUCTURALLY LINKED to each other  induced
conformational change in 1 subunit elicits change in remaining subunits
 o When binding of substrate B TO substrate A:
 INCREASES = positive
OR
 DECREASES = negative

 INHIBITORS  DECREASE velocity of catalyzed rxn

MNEMONIC: EFFECTS OF REVERSIBLE ENZYME INHIBITORS

KoMpetitive INhibition = KM INcrease (Vmax unchanged)

Non-KoMpetitive INhibitor = NO KM INcrease (Vmax decreased)

Uncompetitive Inhibition = BOTH Km & Vmax DECREASE

o 4 TYPES OF REVERSIBLE INHIBITION:

 1) COMPETITIVE INHIBITION
 Reversible inhibitor  binds to active site
 Substrate + Inhibitor COMPETE for active site
 GREATER [S] overcomes inhibition
 Structurally similar to substrate
 CANNOT bind to enzyme @ same time as the substrate
 Can ONLY bind if substrate NOT bound
 CAN be overcome by ADDING MORE SUBSTRATE 
o [S] >> inhibitor
 Bind to free enzyme (=E) ONLY forming an unreactive enzyme-
inhibitor complex (=EI)

 2) NON-COMPETITIVE INHIBITION
 Reversible binding  inhibitor reversibly binds to enzyme
outside of active site to deactivate it
 BOTH inhibitor & substrate BOTH bind to enzyme @ different
spots
 Do NOT compete w/ substrate in binding to active site
  Bind to enzyme @ ALLOSTERIC SITE (NOT active site)
 Bind to free enzyme (=E) OR enzyme-substrate complex (=ES)
WITH the same AFFINITY for BOTH
o  forming an unreactive enzyme-inhibitor complex
(=EI) OR enzyme-inhibitor-substrate complex (=EIS)
 Can bind whether OR not substrate has bound
 CANNOT be overcome by adding more substrate
o Enzymes regain function when inhibitor removed from
system
 Non-competitive inhibitors bind EQUALLY well to enzyme &
enzyme-substrate complex  unlike MIXED INHIBITORS

 3) UNCOMPETITIVE INHIBITION
 Bind to enzyme-substrate complexes (=ES)
o  ONLY forming an unreactive enzyme-inhibitor
complex (=EI)
o INCREASES affinity b/w enzyme & substrate
o REQUIRES preassembled ES complexes
 Formed upon binding  MUST bind @
ALLOSTERIC SITE
 ES complex formation creates
conformational change  allows
uncompetitive inhibitor to bind
o MORE effective when [S] = HIGH

 4) MIXED
 Bind to enzyme @ ALLOSTERIC SITE (NOT active site)
 Bind to free enzyme (=E) OR enzyme-substrate complex (=ES)
BUT has different AFFINITY for each
 o  forming an unreactive enzyme-inhibitor complex
(=EI) OR enzyme-inhibitor-substrate complex (=EIS)

 KCAT = enzyme turnover

number
o How many
substrates [S] an
[E] can turn into
product (P) per 1
second @ the
maximum
speed
o Units: sec-1
 Vmax = maximum speed
of the rxn
 [E]T = total [ ] of
available enzyme

 Hill coefficient = measure of cooperativity

o Ex: Hb  where you see sigmoidal kinetic curves that do not follow Michalis-
Menten

o n<1
 = Negative cooperativity
 o n=1
 NO cooperativity
 Normal Michalis-Menten kinetics
o n>1
 Positive cooperativity

 POSITIVE COOPERATIVITY (Sigmoidal Kinetics)

o Binding of 1 substrate INCREASES affinity for another molecule to bind ** (most
common)

 NEGATIVE COOPERATIVITY
o Binding of 1 substrate DECREASES affinity for another molecule to bind

RXN RATES OF ENZYMES

 1:1:1:1 ratio  because A + B are consumed and C + D are produced @ the same time
 Elementary rxn = rxn that takes place in a single step  in any elementary rxn we can
use the coefficient of that reactant to determine what the order of what that reactant is
within that rate law

 ARRHENIUS EQUATION  rate constant

o = rate constant is independent of the reactant and product concentration
o changing the activation energy (E)  changes K (constant)
 Because enzymes do NOT affect the concentrations of products or
reactants so if enzymes increase rate of rxn and CANNOT increase the
reactants or products they have to increase K (rate constant) value
 If DECREASE EA (activation energy)  INCREASE rate constant and
that is what INCREASES the rate of rxn
 Inside our body the TEMP. = CONSTANT  ALL the rxns take place @
constant temp.  enzymes have to affect the Ea to increase the rate of that
rxn
o A = frequency factor  frequency of collision b/w the reactants
 For a rxn to take place  reactants have to collide w/ a great enough
energy
 A = collision frequency
 If collisions are more frequent  more likely to create products
 INCREASE A value  INCREASE K value
 SMALLER space = more likely to have collisions b/w reactants
 Enzymes can DECREASE space in which the reactants are
colliding  enzymes can INCREASE A because they
DECREASE the space in which reactants are actually allowed to
move
 If enzymes collide MORE the A INCREASES
 DECREASE Ea and INCREASE A  BOTH INCREASE K
(rate constant)  INCREASES the rate of the rxn
 ORDER OF RXN
o = depends on [reactants]
o describes how the rate of rxn depends on [reactants]

MICHAELIS MENTEN
 Constant  reflects the [ ] @ which the rxn rate is @ ½ its max value

 FIRST-ORDER KINETICS
o When [S] = LOW  Vo is proportional to [S]  the amt. of substrate limits the
rate of the rxn (linear)
o When a rxn’s rate is directly proportional to the [reactant]
o exponent = 1  1st order rxn  rate law or rate of rxn depends DIRECTLY
(proportional) to the [ ]
 Double [A] = double the rate
 ZERO-ORDER KINETICS
o When [S] = HIGH  Vo reaches & asymptote (Vmax) and becomes independent
of [S]  adding more substrate will NOT speed up the rxn rate because the
enzymes are already working as hard as they can
 SECOND-ORDER KINETICS
o Bimolecular rxns
o if A = 1st order & B = 1st order
o A+BC
 V = k [A][B]
o By doubling A  double V as long as everything else is constant
o By doubling A & B while K = constant  quadruple V

KINETICS vs. THERMO

 Thermodynamics = predicts:
  Direction
 “Driving force”
 2 forms of energy:
o 1) Heat Energy  movement of molecules
o 2) Potential Energy  energy stored in chemical bonds
 Kinetics = predicts:
 Speed (rate)
 ALL spontaneous processes  proceed toward “states” (macrostates) w/ the
greatest # of accessible mircostates

 Available microstate describes specific detailed microscopic configuration:  that a

system can visit in the course of its fluctuations
 Molecular rotations
 Translations
 Vibrations
 Electronic configuration
 Macrostate describes macroscopic properties such as:
 Temperature
 Pressure

 For gas, liquid, OR solid, the # of available microstates increases w/ T  when

you heat anything  you INCREASE the # of available microstates
 When a liquid vaporizes  the # of available microstates INCREASES
 When a liquid freezes  the # of available microstates DECREASES
NONSPONTANEOUS PROCESS

 There are MORE

available microstates on the right side than on the left side
 If a system gains degrees of freedom  GAINS ENTROPY
 More constituents
 More room to move
 More available quantum states
 More available rotational, vibrational, translational, or electronic states

 SPONTANEOUS PROCESS = INCREASES entropy

LAWS OF THERMODYNAMICS

 1st Law of Thermodynamics

 The change in the internal energy of a system is equal to the work done on it + the heat
transferred to it
 The Law of Conservation of Energy

 In any process  total energy of the universe remains

unchanged: ENERGY is conserved
 A process and its reverse are equally allows by the 1st law

 2nd Law of Thermodynamics

 For a spontaneous process – Entropy – of the universe  meaning the system + its
surroundings – INCREASES ∆ S universe are spontaneous

 Processes that INCREASE

 3rd Law of Thermodynamics
 In any thermodynamic process involving ONLY pure phases @ equilibrium, the entropy
change ∆, S, approaches zero @ absolute zero temperature  also
entropy of crystalline substance approaches 0 @ 0K
  ∆Ssur = depends on the direction of heat flow
 Magnitude of ∆Ssur depends on temperature

** This is ∆H of the system

______________________________________________________________________________

CARBOHYDRATES = Cn(H20)n  carbo + H20

 EASY/ACCESSIBLE energy
 Stacked easy = useful for energy storage
 Carbohydrate & Fats BOTH  HIGH [C-H bonds] = LARGE amt. of energy
STORAGE
 Structure: For each C = 1 O & 2 H’s
 Made of:
 1) C  ratio of 1 C:1 H20 molecule
 Alcohols = present along carbon chain
 2) H
 3) N
o OH (alcohol) on each C EXCEPT for 1  which is EITHER aldehyde OR
ketone attached in straight chain form
o Ring formed when  OH group on chiral C FAR from carbonyl acts as a
nucleophile  ATTACKING carbonyl in nucleophilic addition rxn  forming
hemiacetal
 In GLUCOSE = it is OH on C5
 In AQUEOUS solution = carbohydrates exist = RING form 
equilibrium allows small amt. chain form

o Join together through  DEHYDRATION RXN forming  long chain

POLYSACCHARIDES = energy storage
o REVERSE HYDROLYSIS rxn allows  release of single sugar molecules =
MONOSACCHARIDES

 NAMING:
o 1st  aldo vs. keto
 Depends on whether aldehyde OR ketone present

o 2nd  # in chain
 Glucose = Aldohexose

o C #’ing  begins w/ MOST oxidized end of chain = aldehyde/ketone end

 o Most referred to by common name

o 1) Sugars
o 2) Monosaccharides
o 3) Disaccharides
o 4) Polysaccharides

 EX
o 6C monosaccharides:
 1) Glucose
 2) Galactose
 3) Fructose  BUT fructose = forms 5C ring
o Others:
 1) Ribose
 2) Glyceraldehyde

Prefix:
o DEOXY = -H replaces -OH
o D/L = absolute configuration  assigned based on chirality of C FURTHEST
from carbonyl groups
 Absolute Configuration

 CHIRAL C = furthest from carbonyl group

determines absolute configuration L OR D –
sugar
 FISCHER PROJECTION
o  If OH group on chiral C
FURTHEST from carbonyl = LEFT
= “L”  = RIGHT = “D”

 L & D = ENANTIOMERS
o NOT epimers

o α/β = ANOMERIC configuration

 Classified based on STEREOCHEMISTRY  D OR L

 Classification:
o Aldose, Ketose, Pyranose, Furanose, #ose = sugar w/ # C atoms
 In order to be classified as carbohydrate, a molecule MUST have:
o 1) @ least 3 C backbone
o 2) Aldehyde OR Ketone
o 3) @ least 2 hydroxyl groups
CARBOHYDRATES 3 CATEGORIES:
 1) Monosaccharides
 2) Disaccharides
 3) Polysaccharides

MONOSACCHARIDES = single monomers

 Undergo INTERmolecular nucleophilic substitution rxns w/ monosaccharides  form

ACETALS
o  mechanism monosaccharides use to join & form diasaccharides

HYDROLYSIS OF GLYCOSIDE LINKAGE

 Glycoside linkage = acetal linkage  COVALENT formed in dehydration rxn
o 1) = linkage involving hydroxyl group of anomeric carbon
o 2) = linkage B/W 2 sugars & base of nucleotides
 C1 joins C4 (sometimes C2 or C6)
 Once joined  sugar NO longer free to mutarotate to α ∨β
configuration
 Exs. of glycosidic linkages:
o 1) Starch
o 2) Glycogen
o 3) Nucleotide

STEREOCHEMISTRY
 Single bond = FREE to rotate
 Double bond = LOCKED in place

 ISOMERS = molecules share SAME molecular formula (3 types of isomers)

o 1) Structural Isomers
o 2) Stereoisomers
 Enantiomers
 Diastereomers
o 3) Conformational/Conformer Isomers

 ENANTIOMERS = NON-SUPERIMPOSABLE (NON-idential) MIRROR IMAGES

o OPPOSITE stereochemistry @ EVERY chiral center
o SAME molecular formula
o SAME connectivity
o NOT same molecule  DIFFERENT configuration
o Mirror Images
  SAME properties as 1 another EXCEPT rotation of plane polarized
light  if 1 enantiomer (+) OR ‘D’ then other (-) ‘L’
 o If have EQUAL [ ]’s of 2 ENANTIOMERS  NOT ALL chiral centers
(rotations being cancelled out)

 MESO COMPOUNDS
o Molecules w/ chiral centers = asymmetric C compound, attached to 4
different groups)  BUT have INTERNAL plane of symmetry
 NO optical activity