MJF College of Veterinary & Animal

Science, Chomu (Jaipur)

Department of Vety. Physiology & Biochemistry

7.Biochemistry of Nucleic Acids

Dr. Mahipal Singh Nathawat

Dr. Sagar Kumar Meena
(Assistant Professor )
Vety. Biochemistry
 Introduction
Nucleotides have a variety of roles in cellular metabolism and they are the
constituents of nucleic acids: deoxyribonucleic acid (DNA) and ribonucleic acid
(RNA), the molecular repositories of genetic information. The ability to store and
transmit genetic information from one generation to the next is a fundamental
condition for life.
DNA was discovered in 1869 by Johann Friedrich Miescher, a Swiss researcher.
The demonstration that DNA contained genetic information was first made in
1944, by Avery, Macleod and MacCary.
A segment of a DNA molecule that contains the information required for the
synthesis of a functional biological product, whether protein or RNA, is referred to
as a gene. A cell typically has many thousands of genes, and DNA molecules, not
surprisingly, tend to be very large. The storage and transmission of biological
information are the only known functions of DNA.
RNAs have a broader range of functions, and several classes are found in cells.
Ribosomal RNAs (rRNAs) are components of ribosomes, the complexes that
carry out the synthesis of proteins. Messenger RNAs (mRNAs) are intermediaries,
carrying genetic information from one or a few genes to a ribosome, where the
corresponding proteins can be synthesized. Transfer RNAs (tRNAs) are adapter
molecules that faithfully translate the information in mRNA into a specific
sequence of amino acids.
 Components of nucleic acids
Nucleic acids are the polymers of nucleotides
(polynucleotides) held by 3' and 5' phosphate
bridges.
Nucleotides have three characteristic components:
(1) a nitrogenous (nitrogen-containing) base
(2) a pentose, and
(3) a phosphate
The molecule without the phosphate group is called
a nucleoside. The nitrogenous bases are derivatives
of two parent compounds, pyrimidine and purine.
The base of a nucleotide is joined covalently (at
N-1 of pyrimidines and N-9 of purines) in an N-β
glycosyl bond to the 1 carbon of the pentose,
and the phosphate is esterified to the 5 carbon.
Both DNA and RNA contain two major purine bases, adenine (A) and guanine
(G), and two major pyrimidines.
In both DNA and RNA one of the pyrimidines is cytosine (C), but the second
major pyrimidine is not the same in both: it is thymine (T) in DNA and uracil
(U) in RNA. Only rarely does thymine occur in RNA or uracil in DNA.
 Biologically important purines
The bases such as hypoxanthine, xanthine and uric acid
are present in the free state in the cells and biogically
important purines. The first two are the intermediates in
purine synthesis while uric acid is the end product of
purine degradation.
Purine bases of plants:- Caffeine (of coffee), theophylline
(of tea) and theobromine (of cocoa).
 PURINE, PYRIMIDINE AND NUCLEOTIDE ANALOGS
1. Allopurinol - is used in the treatment of hyperuricemia and gout.
2.5-Fluorouracil, 6-mercaptopurine, 8-azaguanine, 3-deoxyuridine, 5- or 6-
azauridine, 5- or 6-azacytidine and 5-idouracil are employed in the treatment
of cancers.
3.Azathioprine (which gets degraded to 6-mercaptopurine) is used to
suppress immunological rejection during transplantation.
4. Arabinosyladenine - is used for the treatment of neurological disease, viral
encephalitis.
5. Arabinosylcytosine – is used in cancer therapy.
6. Zidovudine or AZT (3-azido 2',3' -dideoxythymidine) and didanosine (di-
deoxyinosine) are sugar modified synthetic nucleotide analogs.
 Structure of DNA
DNA is a polymer of deoxyribonucleotides (or simply deoxynucleotides). It is
composed of monomeric units namely deoxyadenylate (dAMP),
deoxyguanylate (dGMP), deoxycytidylate (dCMP) and deoxythymidylate
(dTMP).
Chargaff's rule of DNA composition- He observed that in all the species he
studied, DNA had equal numbers of adenine and thymine residues (A = T) and
equal numbers of guanine and cytosine residues (G = C). This is known as
Chargaff's rule of molar equivalence between the purines and pyrimidines in
DNA structure.
Single-stranded DNA, and RNAs which are usually single-stranded, do not
obey Chargaff's rule.
DNA DOUBLE HELIX:-
The double helical structure of DNA was proposed by lames Watson and
Francis Crick in 1953 (Nobel Prize, 1962). The structure of DNA double helix is
comparable to a twisted ladder.
Fig.- (A) Watson-Crick model of DNA helix
(B) Complementary base pairing in DNA helix.
Fig.- Phosphodiester linkages in the covalent backbone of DNA and RNA.
Fig.- Hydrogen-bonding patterns in the base pairs defined by
Watson and Crick
 The salient features of Watson-Crick model of DNA
1.The DNA is a right handed double helix. It consists of two
polydeoxyribonucleotide chains (strands) twisted around each other
on a common axis.
2.The two strands are antiparallel, i.e., one strand runs in the 5' to 3'
direction while the other in 3' to 5'direction. This is comparable to two
parallel adjacent roads carrying traffic in opposite direction.
3. The width (or diameter) of a double helix is 20 A° (2 nm).
4. Each turn (pitch) of the helix is 34 A° (3.4 nm) with 10 pairs of
nucleotides, each pair placed at a distance of about 3.4 A°.
5.Each strand of DNA has a hydrophilic deoxyribose phosphate
backbone ( 3'-5' phosphodiester bonds) on the outside (periphery) of
the molecule while the hydrophobic bases are stacked inside (core).
6.The two polynucleotide chains are not identical but complementary
to each other due to base pairing.
7.The two strands are held together by hydrogen bonds
formed by complementary base pairs. The A-T pair has 2
hydrogen bonds while G-C pair has 3 hydrogen bonds. The G =
C is stronger by about 50% than A=T.
8.The hydrogen bonds are formed between a purine and a
pyrimidine only. lf two purines face each other, they would
not fit into the allowable space. And two pyrimidines would
be too far to form hydrogen bonds. The only base
arrangement possible in DNA structure, from spatial
considerations is A-T, T-A, G-C and C-G.
9. The complementary base pairing in DNA helix proves
Chargaffs rule. The content of adenine equals to that of
thymine (A = T) and guanine equals to that of cytosine (G = C).
10. The genetic information resides on one of
the two strands known as template strand or
sense strand. The opposite strand is antisense
strand.
The double helix has (wide) major grooves and
(narrow) minor grooves along the
phosphodiester backbone.
 Different Conformations of DNA Double Helix
Characteristics A-DNA B-DNA C-DNA Z-DNA

Conditions 75 % relative humidity; 92 % relative humidity; 60 % relative humidity; Very high salt
Na+, K+, Cs+ions Low ion strength Li+ions concentration

Shape Broadest Intermediate Narrow Narrowest

Helix sense Right -handed Right -handed Right -handed Left-handed
Helix diameter 25.5 A◦ 23.7 A◦ 19.0 A◦ 18.4 A◦
Rise per base pair (A°) 2.3 A◦ 3.4 A◦ 3.32 A◦ 3.8 A◦

Base pairs per turn of helix 11 10.4 9.33 12 (=6 dimers)

(‘n’)

Helix pitch 25.30 A◦ 35.36 A◦ 30.97 A◦ 45.60 A◦

Rotation per base pair
+32.72 A◦ +34.61 A◦ +38.58 A◦ -60◦ (per dimer)
Base pair tilt 19◦ 1◦ 7.8◦ 9◦

Major groove Narrow and very deep Wide and quite deep - Flat

Minor groove Very broad and shallow Narrow and quite deep - Very narrow and deep
 OTHER TYPES OF DNA STRUCTURE
Bent DNA:-
Bending in DNA structure has been reported due to photochemical damage or
mispairing of bases.
Certain antitumor drugs (e.g. cisplatin) produce bent structure in DNA. Such
changed structure can take up proteins that damage the DNA.
Triple-stranded DNA:-
Triple-stranded DNA formation may occur due to additional hydrogen bonds
between the bases. Thus, a thymine can selectively form two Hoogsteen
hydrogen bonds (non-Watson-Crick pairing) to the adenine of A-T pair to form
T-A-T and cytosine can also form two hydrogen bonds with guanine of G-C
pairs that results in C-G-C.
Four-stranded DNA:-
Polynucleotides with very high contents of guanine can form a novel tetrameric
structure called G-quartets. These structures are planar and are connected by
Hoogsteen hydrogen bonds.
 (b) C (i) C (ii)

(a) Base-pairing patterns in one well-characterized form of triplex DNA. The Hoogsteen
pair in each case is shown.
(b) An outline of Hoogsteen triple helical structure of DNA.
(C) Four-stranded DNAs tructure (i) Parallel G-tetraplex (ii) Antiparallel G-tetraplex
 Denaturation of DNA Strands
The two strands of DNA helix are held together by hydrogen bonds.
Disruption of hydrogen bonds (by changes in pH or increase in temperature)
results in the separation of polynucleotide strands. This phenomenon of loss
of helical structure of DNA is known as denaturation.
Renaturation (or Reannealing) is the process in which the separated
complementary DNA strands can form a double helix.
The phosphodiester bonds are not broken by denaturation. Loss of helical
structure can be measured by increase in absorbance at 260 nm (in a
spectrophotometer).

Fig.- Diagrammatic representation of denaturation and renaturation

of DNA.
Melting temperature (Tm) is defined as the temperature at
which half of the helical structure of DNA is lost.
Since G-C base pairs are more stable (due to 3 hydrogen bonds)
than A-T base pairs (2 hydrogen bonds), the Tm is greater for
DNAs with higher G-C content.
Thus, the Tm is 65°C for 35 % G-C content while it is 70°C for 50
% G-C content.
 Oganization of DNA in the Cell
The double-stranded DNA helix in each chromosome has a length that
is thousands times the diameter of the nucleus. In humans, a 2-meter
long DNA is packed in a nucleus of about 10 µm diameter.
This is made possible by a compact and marvellous packaging, and
organization of DNA inside in cell.
In prokaryotic cells, the bacterial chromosomes are packed in the form
of nucleoids, by interaction with proteins and certain cations
(polyamines).
In eukaryotic cells, the DNA is associated with various proteins to form
chromatin which then gets organized info compact structures namely
chromosome.
The DNA double helix is wrapped around the core proteins namely
histones which are basic in nature. The core is composed of two
molecules of histones (H2A, H2B, H3 and H4). Each core with two turns
of DNA wrapped round it is termed as a nucleosome, the basic unit of
chromatin.
 RNA vs. DNA
RNA is a polymer of ribonucleotides held together by 3',5’-
phosphodiester bridges.
RNAs have specific differences from DNA-
1.Pentose:- The sugar in RNA is ribose in contrast to deoxyribose in
DNA.
2.Pyrimidine:- RNA contains the pyrimidine uracil in place of thymine
(in DNA).
3. Single strand:- RNA is usually a single stranded polynucleotide.
4. Chargaff's rule-not obeyed:-Due to the single-stranded nature, there
is no specific relation between purine and pyrimidine contents.
5. Susceptibility to alkali hydrolysis:- DNA cannot be subjected to alkali
hydrolysis due to lack of hydroxyl group.
6.Orcinol colour reaction:- RNAs can be histologically identified by
orcinol colour reaction due to the presence of ribose.
ATP is the
FUNCTIONS OF NUCLEOTIDES
principal form of chemical energy, available to cells. It is used as a
phosphorylating agent and is also involved in muscle contraction, active transport and
maintenance of ionic gradients.
Nucleotides are the monomeric units of nucleic acids (DNA & RNA).
Nucleotides are used in the synthesis of second messengers like cAMP and cGMP for the
hormonal functions.
Many of the regulated steps of metabolic pathways are controlled by intracellular
concentrations of nucleotides.
They serve as a carrier of high-energy intermediates in the biosynthesis of carbohydrates,
lipids and proteins. e.g.
– GTP is involved in the synthesis of glucose (gluconeogenesis).
– GDP is involved in the oxidation of α - Ketoglutaric acid to succinyl CoA to form
GTP.
– Uracil derivatives UTP is involved in the synthesis of glycogen and also in the
epimerization of galactose and glucose (lactose biosynthesis).
– Cytosine derivatives are involved in the biosynthesis of phosphoglycerides in
animal tissues.
Nucleotides are also structural components of several coenzymes of B complex vitamins.
e.g. NAD, FAD, and pantothenic acid in Co- enzyme A.
Biologically important nucleoside, S- adenosylmethionine (adenosyl derivative) is involved
in several transmethylation processes.
 Types of RNA & their functions
The three major types of RNAs-
1. Messenger RNA (mRNA): 5-10 %
2. Transfer RNA (tRNA): 10-20 %
3. Ribosomal RNA (rRNA): 50-80 %
Messenger RNA (mRNA)-
The eukaryotic mRNA is capped at the 5'-terminal end by 7-
methylguanosine triphosphate and this cap helps to prevent the
hydrolysis of mRNA by 5'-exonucleases and involved in the
recognition of mRNA for protein synthesis.
The 3'-terminal end of mRNA contains a polymer of adenylate
residues (20-250 nucleotides) which is known as poly (A) tail.
This tail may provide stability to mRNA and preventing it from
the attack of 3'-exonucleases.
Transfer RNA (tRNA)-
Transfer RNA (soluhle RNA) molecule contains 71-80 nucleotides
(mostly 75) with a molecular weight of about 25,000. The
structure of tRNA (for alanine) was first elucidated by Holley.
The structure of tRNA (Clover leaf modal)-

tRNA contains mainly four arms, each arm with a base

paired stem.
1.The acceptor arm:-This arm is capped with a sequence
CCA (5' to 3‘). The amino acid is attached to the acceptor
arm.
2.The anticodon arm:- This arm, with the three specific
nucleotide bases (anticodon), is responsible for the
recognition of triplet codon of mRNA.
3.The D arm:- lt is so named due to the presence of
dihydrouridine.
4.The TYC arm:- This arm contains a sequence of T,
pseudouridine (represented by psi, Y) and C.
5. The variable arm:- This arm is the most variable in tRNA.
Based on this variability, tRNAs are classified into 2
categories:
(a)Class I tRNAs : The most predominant (about 75 %) form
with 3-5 base pairs length.
(b) Class ll tRNAs : They contain 13-20 base pair long arm.
Ribosomal RNA (rRNA)-
The ribosomes are the factories of protein synthesis. The
eukaryotic ribosomes are composed of two major nucleoprotein
complexes-60S subunit and 40S subunit. The 60S subunit
contains 28S rRNA, 5S rRNA and 5.8S rRNA while the 40S subunit
contains 18S rRNA. They play a significant role in the binding of
mRNA to ribosomes and protein synthesis.
CATALYTIC RNAs-RIBOZYMES-
The RNA component of a ribonucleoprotein (RNA in association
with protein) is catalytically active.
Ribonuclease P (RNase P) is a ribozyme containing protein and
RNA component.
It is believed that ribozymes (RNAs) were functioning as catalysts
before the occurrence of protein enzymes, during the course of
evolution.
THANKS