NUCLEIC ACIDS

DR. RIFFAT IQBAL

 NUCLEIC ACIDS

• Nonprotein nitrogenous substances

• Polymers of monomeric units, nucleotides
• Nucleic acids are required for the storage and expression
of genetic information.
• There are two chemically distinct types of nucleic acids:
• Deoxyribonucleic acid (DNA)
• Ribonucleic acid (RNA)
DEOXYRIBONUCLEIC
ACID

• DNA is present in chromosomes, mitochondria and

chloroplast
• Every nucleated cell
• Nuclear DNA is bound to basic proteins called Histones
 DNA

• DNA is a polymer of deoxyribonucleotides covalently

linked by 3′→5′–phosphodiester bonds
• With the exception of a few viruses that contain single-
stranded (ss) DNA, DNA exists as a double-stranded (ds)
molecule, in which the two strands wind around each
other, forming a double helix.
 DNA

• In eukaryotic cells, DNA is associated with various types

of proteins (known collectively as nucleoproteins)
present in the nucleus

• In prokaryotes, the protein–DNA complex is present in a

nonmembrane-bound region known as the nucleoid
Structure of DNA

Primary

Secondary

Higher
organization
 Primary structure of DNA
• Chromosomal DNA consists of very long DNA molecules
• Each DNA molecule is a polymer of about 1010
deoxyribonuleotides
• Number and sequence of different deoxyribonucleotides
in its strands joined together by phosphodiester bonds
• Four different types of deoxyribonucleotides

Adenine Guanine
deoxyribonucleotide deoxyribonucleotide
dA dG

Thymine Cytosine
deoxyribonucleotide deoxyribonucleotide
dT dC
 3′→5′-Phosphodiester bond

• Each nucleotide is linked to neighboring nucleotide through

3′→5′-Phosphodiester bond

• Bond joins the 3'-hydroxyl group of the deoxyribose of one

nucleotide to the 5′-hydroxyl group of the deoxyribose of an
adjacent nucleotide through a phosphate group
 Ends of DNA strand

• Linear polydeoxyribonucleotides strand with 2 free ends

on both sides

• The end with a free phosphate group without

phosphodiester linkage is called 5′-end
• The opposite end bearing a free hydroxyl group is called
3′-end
 Polarity of DNA

• One DNA strand runs from 3’ 5’ is coding/ template

strand
• The opposite strand runs from 5’ 3’ direction…
noncoding strand

• The bases located along the resulting deoxyribose–

phosphate backbone are, by convention, always written
in sequence from the 5′-end of the chain to the 3′-end.
• Sugar –PO4 residues connected by PDB form the
constant part of backbone

• Purine/pyrimidine bases linked to the sugar molecule

located inside the backbone form variable part of the
primary structure of DNA

• Sugar –PO4 stabilize the molecule

• Bases carry the genetic information

Secondary structure of DNA

• It is the double stranded helical shape of DNA formed

by the two polydeoxyribonucleotide strands coiled
around a central axis

• Watson- Crick model in 1953

 Double helix
• PDBs in the two interwoven strands run in opposite
direction

• The 5′-end of one strand is paired with the 3′-end of the

other strand

• The strands are present in antiparallel manner

• In the DNA helix, the hydrophilic deoxyribose–
phosphate backbone of each strand on the outerside
form the walls

• Hydrophobic bases are stacked inside

 Base pairing in double helix

• The bases of one strand of DNA are paired with the bases
of the second strand
▪ A is bonded to T by 2 H – bonds
▪ G is bonded to C by 3 H – bonds

• The base pairs are perpendicular to the long axis of the

helix
• Hydrogen bonds, plus the hydrophobic interactions
between the stacked bases, stabilize the structure of the
double helix
The specific base pairing in DNA leads to the Chargoff ‘s
Rule:

• In any sample of dsDNA, the amount of adenine equals

the amount of thymine, the amount of guanine equals
the amount of cytosine

• Total amount of purines = amount of pyrimidines

• A+G = T+C
 Grooves of DNA

• The spatial relationship between the two strands in the

helix creates a major (wide) groove and a minor (narrow)
groove
 Grooves of DNA

• These grooves provide access for the binding of

regulatory proteins to their specific recognition
sequences along the DNA chain.

• Certain anticancer drugs, such as dactinomycin

(actinomycin D), exert their cytotoxic effect by
intercalating into the narrow groove of the DNA double
helix, thus interfering with DNA and RNA synthesis.
Higher organization of DNA

• Nucleosomes
 Histones and the formation of
nucleosomes
• There are five classes of histones, designated H1, H2A,
H2B, H3, and H4.

• These small proteins are positively charged at physiologic

pH as a result of their high content of lysine and arginine.
Because of their positive charge, they form ionic bonds
with negatively charged DNA.
• Histones, along with positively charged ions such as Mg +2,
help neutralize the negatively charged DNA phosphate
groups.
 Nucleosomes

• Two molecules each of H2A, H2B, H3, and H4 form the

structural core of the individual “nucleosome beads”

• Around this core, a segment of the DNA double helix is

wound nearly twice, forming a negatively supertwisted
helix
• Neighboring nucleosomes are joined by “linker” DNA
approximately fifty base pairs long.

• Histone H1, of which there are several related species, is

not found in the nucleosome core, but instead binds to
the linker DNA chain between the nucleosome beads.
• Nucleosomes are further arranged into increasingly more
complex structures that organize and condense the long
DNA molecules into chromosomes that can be
segregated during cell division

• The complex of DNA and protein found inside the nuclei

of eukaryotic cells is called chromatin
 Separation of the two DNA
strands in the double helix

• The two strands of the double helix separate when

hydrogen bonds between the paired bases are disrupted

• Disruption can occur in the laboratory if the pH of the

DNA solution is altered so that the nucleotide bases
ionize, or if the solution is heated
• Phosphodiester bonds are not broken by such treatment

• PDB between nucleotides (in DNA or RNA) can be

cleaved hydrolytically by chemicals, or hydrolyzed
enzymatically by a family of nucleases:
• deoxyribonucleases for DNA
• ribonucleases for RNA
 Denaturation of DNA

• Complete separation or unwinding of DNA strands in

vitro is called denaturation

• H – Bonds between bases break when DNA is treated

above a certain temperature/ melted
 Melting temperature of DNA

• The temperature at which DNA is half denatured is

known as the melting temperature ,Tm of DNA

• Because there are three hydrogen bonds between G and

C but only two between A and T

• DNA that contains high concentrations of A and T

denatures at a lower temperature than G- and C-rich DNA
 DNA Annealing

• When the separated DNA strands are slowly cooled, the

complementary strands are paired again

• Renaturation /or annealing

• At temperature below Tm
 Structural forms of DNA

• There are three major structural forms of DNA as

described by Watson and Crick in 1953:
▪ A- DNA
▪ B- DNA
▪ Z- DNA
 B- DNA

• The B form is a right-handed helix with ten residues per

360° turn of the helix, and with the planes of the bases
perpendicular to the helical axis

• Chromosomal DNA is thought to consist primarily of B-

DNA
 A - DNA

• It is also a right-handed helix, but there are eleven base

pairs per turn, and the planes of the base pairs are tilted
20° away from the perpendicular to the helical axis

• Not present under normal physiological conditions

 Z - DNA

• Z-DNA is a left-handed helix that contains about twelve

base pairs per turn

• Stretches of Z-DNA can occur naturally in regions of DNA

that have a sequence of alternating purines and
pyrimidines
 Circular DNA molecules

• Each chromosome in the nucleus of a eukaryote contains

one long linear molecule of dsDNA, which is bound to a
complex mixture of proteins to form chromatin.

• Eukaryotes have closed circular DNA molecules in their

mitochondria, as do plant chloroplasts.
• Most species of bacteria also contain small, circular,
extrachromosomal DNA molecules called plasmids

• Plasmid DNA carries genetic information, and undergoes

replication that may or may not be synchronized to
chromosomal division
• Plasmids may carry genes that convey antibiotic
resistance to the host bacterium, and may facilitate the
transfer of genetic information from one bacterium to
another

• Plasmids are used as vectors in recombinant DNA

technology
RIBONUCLEIC ACID
 RNA

• The genetic master plan of an organism is contained in

the sequence of deoxyribonucleotides in its DNA

• ribonucleic acid (RNA)—the “working copies” of the

DNA—through which the master plan is executed
• The copying process, during which a DNA strand serves
as a template for the synthesis of RNA, is called
transcription

• The final product of gene expression, therefore, can be

RNA or protein, depending upon the gene
 RNA

• Polymer of ribonucleotides
• A, G, C, U
• T absent
• Phosphodiester bonds
• D – Ribose
• Nucleolus, Nissl granules, ribosomes, mitochondria and
cytoplasm
Structure of RNA
 Primary structure of DNA

• Number and sequence of ribonucleotides in linear strand

• 3′→5′-Phosphodiester bond joins the 3'-hydroxyl of one

ribose to the 5′-hydroxyl of adjacent ribonucleotide
 Secondary structure of RNA
• Coil formation by various polyribonucleotides

• Intra-chain H – bonds between

• G – C (3 H bonds)
• A – U (2 H bond with N3 and C4 of uracil)

• Hydrophobic interactions between purine & pyrimidine

bases stabilize the coiled structure
 Tertiary structure of RNA

• Folding of RNA molecule into 3D structure

• Compact coiled globular structure is formed due to

▪ Cross linking at various sites
▪ H – bonds
▪ Hydrophobic interactions
 Types of RNA

mRNA rRNA tRNA

Sn-RNA Hn-RNA
 Messenger RNA

• Most heterogeneous w.r.t size & stability

• Mol. wgh varies from 3 x 104 – 2 x 106

• 103 – 104 ribonucleotides per molecule

• Major bases:
• A, G, C, and U
 mRNA

• Minor bases:
▪ Methylpurine
▪ Methylpyrimidine

• Formed from DNA template strand during transcription

 mRNA

• CODON: mRNA carries specific nucleotide sequences in

the form of a set of three bases “triplets”

➢Codons are responsible for synthesis of a specific

protein molecule
 Ends of mRNA
• “POLY A TAIL”: 3´-OH end carries a polymer of
adenylate ribonucleotides containing 20-250 residues
▪ Maintain intracellular stability of specific mRNA by
preventing attack of 3´-exonucleases

• 5´-OH end carries a cap structure (7 methylguanosine

triphosphate)
▪ Recognition of protein synthesis site
▪ Prevents attack of 5´-exonucleases
 Ribosomal RNA
• Makes up about 80% percent of the total cellular RNA

• rRNAs are found in association with several proteins as

components of the ribosomes

• Sedimentation coefficient is 80S

• Two subunits:
▪ Larger one 60S
▪ Smaller one 40S
 Transfer RNA

• Smallest of the three major species of RNA molecules

• Single stranded, globular

• About 15% of total RNA in the cell

• They remain largely in cytoplasm

 Primary structure of tRNA

• Single strand held by PDB

• Consists of approximately 75 ribonucleotides

• Bases are:
• A, G, C, U and pseudouridine (ψ)
 Secondary structure of tRNA

• Each single strand is folded to form a clover-leaf

secondary structure

• Folds are stabilized by H-bonds b/w complementary

bases in different portions of the same strand

• These double stranded helical structures are called as

stems
• All t-RNA molecules contain 4 main arms or loops
1. Acceptor Arm
• Unpaired sequences of Cytosine-cytosine-Adenine (C-
C-A) at the 3’ end also called as acceptor end

• The 3’ – OH terminal of adenine may bind with the α-

COOH of a specific amino acid and carry the latter as an
aminoacyl t-RNA complex to ribosomes for protein
synthesis

• Borne by base-paired acceptor stem, whose bases are H-

bonded with last few bases at the 5’ end of tRNA
2. Anticodon Arm
• This is another unpaired and non-bonded loop carrying
specific sequences of three bases constituting the
anticodon
• The bases of anticodon are hydrogen bonded with three
complementary bases of codon of m-RNA
• The base pair stem leading to anticodon loop is called
the anticodon stem
3. D-arm:
The third is the D-arm because it contains the
base dihydrouridine

4. TψC arm:
Contains thymine, pseudouridine and cytosine

5. Variable arm or extra arm:

Extra arm is most variable structure of t-RNA and it forms
the basis of its classification
 Classification of tRNA

(a) Class I tRNA:

• About 45 per cent of all t-RNA belong to this class and
have 3-5 base pairs in its extra arm, e.g. Ala-t-RNA.

(b) Class II tRNA:

• This forms about 25 per cent of total tRNA
• 1321 base pairs in a long chain, e.g. Phe-tRNA.
• The secondary structure of t-RNA is mainly maintained
by the base pairing in these arms and this is a consistent
feature
 Tertiary structure of tRNA
• L- shaped

• Formed by further folding of clover leaf due to H-bonds

b/w T and D arms

• Base paired double helical stems get arranged into two

double helical columns perpendicular to one another

• Locates the aminoacid acceptor and the anticodon

 Small nuclear RNA (Sn-RNA)
• 90-300 nucleotides

• Nucleoplasm, nucleolus, perichromatic granules &

cytoplasm

• Not involved in protein synthesis but processing

• Architecture of RNA
 Heterogenous nuclear RNA (hnRNA)

• Nucleolus

• Large precursor of mRNA

RNAs differ as a group from DNA in several ways:
• They are smaller than DNA

• They contain ribose instead of deoxyribose

• They contain Uracil instead of thymine

• Most RNAs exist as single strands that are capable of

folding into complex structures