Lecture 1-History of Microbiology

- Spontaneous growth debate

o Do microbes come from non-living matter?
o Louis Pasteur neck flask experiment
 Proved that microbes do not come from non-living matter
 Showed that microbes are a bit similar to plants and animals
 Many industrial processes rely on microbial metabolism
 Ex. wine making
- Germ Theory
- Koch’s Postulates
o Able to link specific microbe with a disease
o Rules
 1. Pathogen must be present in all patients with the disease
 2. Pathogen must be isolated and grown in pure culture
 3. Cells from pure culture must be put into healthy animal and cause the same
disease
 4. Suspected pathogen must be reisolated and be the same as the pathogen
from the original
- Enrichment cultures
o Put small amount of environmental sample into selective culture medium
o When organisms grow, take small amount to another container of selective medium and
repeat
o Eventually, isolation of a species can be obtained via streaking
o Specific conditions favor specific organisms

Lecture 2-Bacterial Growth and its Measurement

- Counting bacteria
o Direct microscopic count
 Use counting chamber with rids
 Count number of bacteria per volume
 Problems
 Can’t tell between live or dead bacteria
 Lack of bacterial motility
o Viable count
 Count number of cells capable of forming a colony on solid medium
 Problems
 Takes time for culture to grow
 Can only count living cells
 If counting species on environmental sample, medium may be selective
for some species
- Serial dilutions
o Help separate bacteria if they are too densely populated in original sample
o 10-fold serial dilutions before plating
  Multiple plate count by dilution factor to obtain original number of bacteria
 Ex. if I count bacteria on plate from 10^-6 dilution, I would multiply the
number of CFUs I counted by 10^6
- Turbidity
o OD proportional to number of cells present
o Indirect measurement of bacteria population
o Less light transmitted (more cloudy), the more bacteria there are
o Very fast method
o Data must be graphed
o Problems
 Can’t tell if all same bacterial species
 Doesn’t distinguish between alive and dead cells
- Culture media
o Rich/complex-not chemically defined, contain complex organic molecules
o Defined/minimal media-chemically defined, contains precise amounts of chemicals
 May have organic molecules
 Usually has H2O, C, N, P,etc
 Total cell yield in culture is determined by limiting nutrient
- Bacterial growth
o Bacteria can go through exponential growth if enough resources are present
o Bacteria double each generation time
o Equation:
 N=N0*2n, N=final cell #, N0=initial cell #, n=# of generations
 n=t/g, g=generation (doubling) time, t=time elapsed during exponential
growth
o Growth phases
 Stationary phase
 Environment conditions limit cell growth
o Ex. lack of nutrient, accumulation of toxic waste product
 No net increase/decrease in cell number
 Energy metabolism can still occur, some biosynthetic processes can
continue
 Lag phase
 Cells are not in most rapid exponential growth rate
 Happens when:
o Stationary phase culture is diluted into fresh medium
o Cells are moved from rich to minimal medium
o Culture is moved to different conditions of temperature or
stress
 Bacteria must readjust to new environment and make proteins needed
to survive there
- C and N
o Most animals use NH4+ or NO3- as N source
 o Oxygen
 Oxic zone- area of tube containing O2
 Obligate aerobes-need O2 as terminal electron acceptor
 Microaerophiles-live in places with low O2
 Aerotolerant anaerobes-neither use O2 nor are harmed by it
 Facultative anaerobes-can use O2 or other strategies when no O2 is present
 Grow better in O2
 Obligate anaerobes-must live in places with no O2
 When O2 is present, ROS can be accidentally made by electron-carrying
molecules
 ROS can damage cells, so enzymes are made that remove ROS
- Optimal growth temperature
o Minimum temperature-membrane gels, slow transport so growth cant happen
o Optimum-enzymatic rxns run at max rate
o Maximum-enzymes denature, membrane collapses, thermal lysis
o Bacteria do not regulate their internal temperature
 Extreme bacteria
 Thermophile- >45C
 Hyperthermophile- >80C
 Psychrophile- <15C
- Adaptations to heat
o Problem-enzymes denature in heat
 Solutions
 Less glycine residues in protein
 Increased ionic bonding between acid/base AAs and their hydrophobic
cores
 Chaperones to help refold proteins
 Solutes to stabilize proteins
o Problem-membranes are too fluid
 Solutions
 Increase amount of saturated fatty acid chains in membrane
- Adaptations to cold
o Problem-Proteins have less thermal motion, so they work slower
 Solution
 proteins are more flexible, so they can move around more even when
the temperature drops
o Problem- membrane fluidity decreases
 Solutions
 increase amount of unsaturated fatty acids in membrane
 increase amount of glycine residues in AAs
o Problem-Ice crystals form and damage cell walls
 Solution
 Cryoprotectants to help prevent ice crystal formation
 - Adaptations to osmolarity
o Hypotonic (hypoosmotic) medium
 Cells usually swell
 Solution
 Have rigid cell wall that can withstand pressure increases
 Mechanosensitive channels that can leak solutes when pressure inside
cell increases
- Hypertonic (hyperosmotic) medium
o Cells shrink
 Solution
 Import or make compatible solutions that increase internal osmolarity
o Ex. sugars

Lecture 3-Bacterial Genomics

- Next Generation Sequencing

o Generates short sequence reads (100 bp) that are assembled into a complete genome
 Computer algorithms find overlaps between short sequence reads
 Contigs-long connected sequences
 Initial output, must be connected into complete genome
o PacBio
 Gap closure-Helps connect contigs to each other
- Annotation
o Process of describing what’s in a genome
o Find open reading frames (ORFs) to identify set of proteins encode by organism
 All ORFs start with sequence ATG (start codon) and end in stop codon
 ORFs that produce proteins usually 200 bp or longer
o Find ribosome binding sites (RBSs) upstream of ORFs
 ORFs with RBSs are more likely to be expressed
o Determine codon bias of each ORG compared to others in the genome
 Multiple codons encode some AAs
 Each organism prefers some codons over others
 ORF with different codon bias (uses different codons) than the rest of the ORFs
in the genome is less likely to make a protein
o Predict tRNAs and rRNAs highly conserved across organisms
- Predicting function of encoded protein
o Happens after an ORF is found and predicted
o BLAST predicted protein sequence
 Helps find similar protein sequences in other organisms
 Existence of homologs makes it more likely that ORF encodes a functional
protein
o Find protein domains (Pfam, CDD) and active sites in sequence (PROSITE, MOTIF)
o Predict protein structure
 SWISS-MODEL-based on known structure
  Alphafold-de novo
o Predict protein’s cellular location (PSORTb)
- Bacterial Genome Properties
o Size from 150kb to 13Mb
o About 1 ORF per kb
o 30% of ORFS in a new genome are “hypothetical”
 Can’t determine what their function
o Genes organized in operons
 2+ ORFS found on same mRNA strand
o Genomic islands
 20-200 kb
 Unlike rest of genome
 Have special functions
 Different codon bias
 Absent in closely related species
 Flanked by direct repeat sequences
 Sign of recombination
 Encode for special processes
 Ex. virulence
 Likely acquired by horizontal gene transfer
- What to do with sequenced genome
o Reverse genetics
 Identify predicted proteins of a certain type in genome, knock out gene for each
one individually
 Observe phenotype
o Comprehensive transcriptional analysis
 Measure expression of every gene simultaneously
 Under different environmental conditions or mutant backgrounds
 Use RNA-seq
 Take all RNA, isolate specific RNA type
 Reverse transcribe RNA into DNA
 Sequence DNA with NGS
 Compare transcriptome from two or more different conditions
o Some genes may be expressed more/less in one environment vs
another
o Predict metabolic pathways
 Use BLAST to search genome for proteins with known enzymatic activities,
connect them in pathways
- 1 gene for every 1000 bases

Lecture 4-Transcriptional Regulatory Mechanisms I

- Central dogma
o Bacterial chromosomes found in cytoplasm
 o Transcription and translation can happen simultaneously
o Bacterial genes encoded in operons
 Proteins transcribed on same mRNA strand
 Translated separately due to having different RBSs
- Control
o Cells control gene expression and enzyme activity
- Transcriptional regulation
o RNA polymerase-transcribes DNA into RNA
 Contains sigma factor
 Binds to specific promoter sequence on gene
o Promoter
 Region of DNA where RNA pol binds to
 Better match of promoter sequence with consensus sequence for a sigma
factor makes stronger promoter
 Poor match yields weaker promoter
o Extra proteins required to help RNA pol bind to promoter
 Cell regulates global gene expression patterns by controlling which sigma
factors are present and active
 Sigma factor must be degraded after use
o Accessory transcription factors
 Activate or repress specific genes
 Activator-turns on transcription
 ex. CRP for lac operon
 Repressor-turns off transcription
 Bind to operator sequence of operon, block downstream gene
transcription
 Corepressor-helps repressor block transcription
o Bind to repressor
o Sometimes required to block transcription
o Can be end products of biosynthetic pathways
 Ex. tryptophan is corepressor of trp operon
 Inducer-activate transcription by inactivating repressor
 Bind to repressor, inactivating it
 Ex. lactose for lac operon
o Catabolite repression
 Allows bacteria to use sugars in sequence
 Glucose always used first
 Diauxic growth curve-lag in growth when glucose is gone
 Cells must make new enzymes to metabolize secondary sugar
 CRP-regulates catabolite repression
 Activator
o Required for transcription of lac, mal, ara operons
 Coactivator is cAMP
 o cAMP binds to CRP, activating it
o When glucose is high, cAMP is low, and vice versa

Lecture 5-Microbial Energetics I

- Metabolism
o Two branches-energy conservation (catabolism) and energy consumption (anabolism)
- Metabolic options for energy conservation
o Chemicals-chemotrophy
 Chemoorganotrophs-use organic compounds (contain C) for energy
 Chemolithotrophs-use inorganic compounds for energy
o Light-phototrophy
 Phototrophs-use light for energy
- Redox rxns
o Electron tower
 Reduction potential EI0-tendency of oxidized substance to accept electrons
 top of tower-lowest EI0, greatest tendency to donate electrons
 many compounds can either be electron donors or acceptors, depending on
what other electron carriers are present
o oxidation half rxn
 electron donor gives up electrons and becomes oxidized
o reduction half rxn
 electron acceptor takes electrons and becomes reduced
o construction of redox rxn: just practice
o Differences in EI0 are expressed as Δ EI0= (EI0 of reduction couple)-( EI0 of oxidation
couple)
 If EI0>0, rxn is favorable in direction written
- Relationship between Δ EI0 and ΔGI0
o ΔGI0=-nF Δ EI0
 n=total number of electrons, F= 96.5 kJ/vmole-
 units are kJ/mole of X oxidized
- How do bacteria store energy they get from redox rxns?
o Proton gradient across cytoplasmic membrane
o High energy compounds that power unfavorable rxns
 Ex. ATP, GTP, PEP
o In catabolism, electrons are extracted and transferred to electron carriers
 Ex. NAD+, FAD, NADP+
 Reduced carriers bring electrons to electron transport chain
- Chemoorgantrophs
o Use organic compound as both electron donor and carbon source
o Aerobic chemoorganotroph- use O2 as electron acceptor
o Some substrates are fully oxidized to CO2 and electrons are used to drive PMF and make
ATP
 CO2 excreted as waste product
 o Cs in other substrates used to build cellular molecules/structures
- Chemolithotrophs
o Use inorganic compounds to get electrons
o Can use O2 or other molecules as electron acceptors
o Often autotrophs, obtain C for cellular molecules from CO2
 Carbon fixation
- Phototrophs
o Get energy from light
o Get electrons from H2O or other compounds
o Get C from CO2 (photoautotrophy) or other organic compounds (photoheterotrophy)

Lecture 6-Fermentation vs Respiration

- Fermentation
o Happens when there is no electron acceptor available for respiration
o No ETC
o ATP made via substrate-level phosphorylation
o Energetically inefficient
 Respiration preferred if given a choice
o Starting compound is only partially oxidized
 Fermentation products- Compounds still containing energy that are excreted
by cell as waste
o NAD+ must be regenerated from NADH
 Occurs when oxidized product is reduced to fermentation product
 NAD+ is needed for other metabolic rxns in the cell
o Most carbon from substrate is excreted as partially reduced waste product
 Only small amount of carbon used in biosynthesis
 Cell must use all carbon substrate for energy production
- Glycolysis
o Glucose oxidized to pyruvate
o Pyruvate can be reduced to make fermentation products or fed into the TCA cycle
 If respiration is possible, go to TCA cycle
 if not, make fermentation products and regenerate NAD+
o Net 2 ATP and 2 NADH made at end of glycolysis
- The Pasteur Effect
o Low net yield of ATP per glucose when fermentation occurs
 2 mol ATP/1 mol glucose
o Cell takes in a lot of glucose when going through fermentation since it’s so energetically
inefficient
o When O2 is added and respiration can occur, cell decreases glucose intake and alcohol
production ceases
- Respiration
o Pyruvate completely oxidized to CO2
o Electrons on NADH and FADH2 are moved to an ETC
 o TCA cycle also generates intermediates of many other biosynthetic pathways
o Generates much more energy per substrate than fermentation due to complete
oxidation of glucose
- Electron Transport Chain
o Series of electron donor and acceptor molecules that ends in a terminal electron
acceptor
 Each electron carrier has reduction potential
o Span cytoplasmic membrane
o Electrons move from lower to higher EI0 as they travel through the complexes
 Energy released as electrons are moved
o Energy released is converted into a proton gradient
o Electron carriers:
 NAD+/NADH
 Carries 2 H+ and 2 electrons
o Involved in other pathways too
 FMN
 Intermediate carrier in ETC
 Carries 2 H+, 2 electrons
 Cytochromes
 Contain heme cofactors
o Fe in heme group cycles between 2+ and 3+ states as electron is
accepted and donated
 Only carry electrons
 FeS clusters
 Covalently bonded to ETC by cysteine residues
o Fe cycles between 2+ and 3+ states as electron is accepted and
donated
 Only carry electrons
 Quinone molecules
 Diffuse in the plane of the cytoplasmic membrane
 Interact with different donor and acceptor complexes
 Carry 2 H+ and 2 electrons
 When accepting electron from an upstream donor, also takes a H+ from
cytoplasm
o When electron is passed to a downstream acceptor, the H+ is
released into the periplasm, helping to generate PMF

Lecture 7-Aerobic and Anaerobic Respiration

- Properties of all ETCs

o Electrons passed from lower to higher reduction potential
o Complexes embedded in cytoplasmic membrane
o Energy released from favorable electron transfers is stored as a proton gradient
o Modular
  Individual complexes can be substituted when bacteria can perform multiple
kinds of respiration
 Depends on the electron donors and acceptors available
- Chemoorganotrophs
o Get electrons from carbon sources using NADH as intermediate carrier
 ETC can start with complex I
 NADH dehydrogenase
o Receives electrons from NADH
 Complex II
 Succinate dehydrogenase
 IIIIV or IIIIIIV
- ETC process
o Electrons are passed between the membrane complexes by quinone molecules in the
cytoplasmic membrane
 Quinone molecules also accept H+ from the cytoplasm to become fully reduced
 When quinone releases its electrons to a downstream carrier, the H+ it had is
released into the periplasm and contributes to PMF
o Cytochrome c can also shuttle electrons between complexes
o End of the ETC
 Electrons given to exogenous acceptor
 Aka terminal electron acceptor
 Reduced acceptor excreted as waste
 Aerobic respiration-O2 is acceptor, H2O is waste product
 Anaerobic respiration-other compounds are reduced by terminal
reductases different from complex IV
- Oxidative Phosphorylation
o Proton gradient generated by electron transport powers ATP synthesis
o Around 3 H+ per ATP produced
- Fermentation ion gradients
o During fermentation, ATP generated by substrate-level phosphorylation runs ATPase
backwards
 ATP is hydrolyzed to make a proton gradient
 Proton gradient powers membrane transport rxns
- Assimilative vs dissimilative reduction
o Assimilative reduction-compounds are reduced to build cellular macromolecules
 Cell only reduces the specific amount needed for growth
o Dissimilative reduction-compounds are reduced for energy conservation
 Large amounts are reduced
 Cell excretes reduced product as waste
- Nitrate reduction
o NO3- can be used as an electron acceptor
o Nitrate reductase complex-used to reduce NO3- in ETC
 Replaces cytochrome oxidase from aerobic respiration
  When NO3- is reduced by nitrate reductase, H+ is not pumped across
cytoplasmic membrane
 Weaker proton gradient, so less ATP is produced from NO3- reduction
o O2 reduction is preferred over NO3- reduction
 Gene expression for NO3- reduction is regulated
 in presence of O2 and NO3-, cells will inhibit transcription of nitrate
reductase gene
- Ferric iron reduction
o Fe3+ can be used as an electron acceptor, but in its natural mineral form it is
inaccessible to cells
o Solutions
 Electron shuttling compounds
 Reduced by cytochromes on cell surface
 Diffuse from cell and transfer electrons to Fe3+ minerals
 Can be obtained in nature or made by cell
o Ex. AQDS, menaquinone
 Direct transfer
 Move electrons from cellular cytochromes right onto Fe3+
 Electrons in quinone pool are transferred on heme groups to series of
cytochromes spanning the inner and outer membranes
o Cytochromes pass electrons onto Fe3+
 Protein Nanowires
 Extend from cell
 Each nanowire contains heme groups that conduct electrons away from
cell and onto Fe3+
- Methanogenesis
o Anaerobic respiration
o Happens only in archaea
 Strict anaerobes
o Electron donor is H2
o Electron acceptor is C in CO2, methanol, or acetate
 Fully reduced to methane, a waste product
o Uses unique one-carbon and electron-carrying cofactors
 MF
 MP
 CoM
 CoB
o Generates Na+ motive force or PMF as one of final steps in methane generation
 Where energy is conserved
 This is the step that makes it respiration!

Lecture 8-Chemolithotrophy

- Chemolithotrophs
 o Metabolize inorganic compounds
o Have alternative ways of obtaining carbon
 Autotrophs-obtain C from CO2
 Mixotrophs-carbon source is organic compound, but not used to obtain
electrons
o ATP synthesis is by oxidative phosphorylation, powered by PMF
o NADH must be generated, regardless of the electron donor being used
 If reduction potential of the electron donor couple is greater than NAD+/NADH,
reverse electron transport must occur
 PMF provides energy for this process
 Electrons are pushed uphill onto NAD+
- Hydrogen as an inorganic electron donor
o Some H2 acceptors are aerobes, others are anaerobes
 Aerobic H2 acceptors can obtain carbon from CO2 via carbon fixation
 Some aerobic H2 oxidizers can also grow as chemoorganotrophs
 Would rather use sugars instead of H2
o Sugars repress genes for H2 oxidation and carbon fixation
o Electron flow in aerobic H2 oxidation
 Membrane-bound hydrogenase gives electrons to quinone pool
 Quinone passes electrons to cytochrome complexes, which give electrons to O2
 PMF is generated, contributing to ATP synthesis
 Cytoplasmic hydrogenase-uses H2 to reduce NAD+ into NADH
 NADH made this way can be used in reduction biosynthetic rxns
- Sulfur as an inorganic electron donor
o Sergei Winogradsky
 Observed bacteria that grew in lots of H2S had granules
 If deprived of H2S, the granules would disappear, but the bacteria would still
grow for a period of time
 These bacteria oxidize H2S to S0
 Energy extracted is used for growth
 S0 stored in granules
o S0 can be used as an electron source once H2S is depleted
- Iron-oxidizing bacteria (FeOB)
o Fe2+/Fe3+ has different EI0 at different pHs
 At pH 7, iron oxidation can be coupled with both O2 and NO3- reduction
 Competing rxn: Fe2+ can be abiotically oxidized by O2 to form Fe3+
o FeOB at pH 7 must live in anoxic or microoxic environments
o Metabolism has to compete with abiotic oxidation of Fe2+
 At pH2, can be coupled with only O2 reduction
 Energetically inefficient
 Large amounts of waste products generated
 1 electron per Fe atom
o Electron flow at low pH
  Fe2+ oxidation is performed by a cytochrome c on the outer membrane
 Fe remains extracellular
 Rusticyanin moves electrons from cytochrome c into complexes in the ETC
 Reverse electron flow occurs-PMF moves electrons to NADH oxidoreductase
that reduces NAD+ to NADH
o Neutral Fe oxidation
 Since Fe oxidation only yields 1 electron per atom, lots of abiotic Fe oxidation
occurs
 Problem: Oxidized iron forms insoluble crystals
 Mariprofundus ferroxydans
o Produces a polysaccharide stalk that extends from its cell body
o Fe3+ minerals precipitate out onto stalk, removing any insoluble
Fe from the cell body

Lecture 8-Phototrophy

- Phototrophs
o Obtain energy from light, but must still obtain electrons from chemical sources
 Electrons used to generate PMF or make FADH2, NADH for redox rxns
- Photosynthesis
o O2 is electron acceptor
o Oxygenic photosynthesis- H2O is electron donor
 O2 produced as waste product
o Anoxygenic photosynthesis-H2S or another reduced chemical is electron donor
 O2 not produced as waste product
o Light and dark rxns
 Light rxn-light energy stored as chemical energy and reducing power
 phototrophy
 Dark rxn-chemical energy and reducing power are used to reduce CO2 into
organic compounds
 Carbon fixation
- Phototrophy
o Light harvested with photosynthetic reaction centers (RC)
 Chlorophyll pigments are bound to proteins or associated with photosynthetic
membranes via a phytol group
 ex. chlorophyll a absorbs red and blue light and reflects green
 ex. bacteriochlorophylls absorb light of all different wavelengths
 Type 1-Photosystem I (PSI)
 First stable electron acceptor is a Fe/S cluster
 Fe/S type RC
 Type 2- Photosystem II (PSII)
 First stable electron acceptor is a quinone molecule
 Q-type RC
 o PSI or PSII can be used individually to generate PMF and reducing power
o Oxygenic photosynthesis uses both PSI and PSII in the same membrane
- Principles of phototrophic transport
o Light excites chlorophyll, allowing it to donate electrons to an electron acceptor to due
decrease in reduction potential
o Electrons move down as reduction potential increases
o PMF is generated
o Reverse electron flow occurs to make NADH if first stable electron acceptor has higher
reduction potential than NAD+
- Electron flow in anoxygenic photosynthesis
o After being excited by light, chlorophyll P870 donates electron to the next acceptor
o Cyclic electron flow-electron is passed through acceptors, until it returns to the original
chlorophyll
 P870Quinone moleculescytochromesP870
 During transfer of electrons from quinone to cytochrome bc1, PMF is generated
 When electrons are removed from the pathway to generate NADH (reverse
electron flow), external electrons must be provided to reduce P870
 Come from electron donors like H2S
o PSI (Fe/S type RC) can also be used, but unknown if cyclic electron flow is utilized
- Electron flow in oxygenic photosynthesis
o Z-scheme
 Uses both PSI and PSII
 Part 1
 LightP680PhQAQBPQ poolcyt b6fplastocyaninP700
 When PQ accepts electrons, it also picks up protons from the
dissociation of water
o When it donates electrons to cyt b6f, protons are pumped into
periplasm to generate PMF
 Part 2
 LightP700chl aFe/S clusterFdFNRPQ pool (cyclic electron
flow)
o Fd can keep electrons for use in other reactions
o In linear electron flow, FNR reduces NADP+ to NADPH for use in
other rxns
o In cyclic electron flow, FNR donates electrons to the PQ pool to
generate more PMF
 No reducing power is made though
- Internal membrane systems
o Used to accommodate all of the photosynthetic proteins, since they span such a huge
area
- Light-harvesting pigments
o Chlorosomes
 Contain tightly packed crystalline array of bacteriochlorophyll (Bch)
 Excitation energy transferred from Bch through intermediate proteins
to rxn centers in the cytoplasmic membrane
o Phycobilisomes (antenna complexes)
 In cyanobacteria
 Absorb light at different wavelengths
 Bacteria can use different phycobilisomes in different environments as a
result
 Absorb light energy and transfer it to rxn center

Lecture 10-Carbon Fixation

- Dark reaction
o Converts CO2 into carbohydrates
o ATP and reducing power generated by phototrophy reduces CO2
o Calvin cycle or Reverse TCA cycle are used
- Calvin cycle
o Uses RuBisCo and PRK enzymes
o Requires NADPH and ATP
o RuBisCo
 Catalyzes conversion of Ribulose bisphosphate to PGA
 Carboxylation
 Competing rxn: Ribulose bisphosphate+O21 PGA + 1 phosphoglycolate (PG)
 PG is toxic to cells
 Mechanisms required to favor carboxylation
o PRK
 Catalyzes conversion of ribulose phosphate to ribulose bisphosphate
- Concentrating carbon
o Problem
 CO2 is natural substrate of Rubisco, but HCO3- is more prevalent
 Can’t accumulate CO2 inside lipid-bound compartment
o Solution
 Carboxysomes
 Cellular compartment that helps concentrate CO2 inside cell
 Protein shell-controls what goes in and out of compartment
 Inside contains high amount of Rubisco and carbonic anhydrase
 Physical, functional and genetic organizations
o Transcribed on operon
o CO2 can’t diffuse out, O2 can’t diffuse in, favoring carboxylation
rxn
 Inducible high-affinity transporters maximize uptake of CO2 and HCO3-
 o CO2 is reduced to HCO3- after import
o HCO3- cannot pass through cytoplasmic membrane
 HCO3- influx into carboxysome powered by Na+ influx
or ATP
 HCO3- + H+CO2+ H2O
- Reverse TCA Cycle
o found in green sulfur bacteria and archaea
o ATP and reduced Fd from light rxn are used to drive TCA cycle in reverse
o ATP citrate lyase is diagnostic for this pathway