Nucleic Acid

Introduction
• There are two types of nucleic acids, namely Deoxyribonucleic
acid (DNA) and Ribonucleic acid (RNA)
• Primarily, nucleic acids serve as repositories and transmitters of
genetic information
• The amino acid sequence of every protein in a cell, and the
nucleotide sequence of every RNA, is specified by a nucleotide
sequence in the cell’s DNA
• 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
• Nucleic acids are the polymers of nucleotides (polynucleotides)
held by 3' and 5' phosphate bridge
INTRODUCTION

Nucleotides and nucleic acids

Nucleotides are the building blocks of nucleic acids

NUCLIEOTIDE
RNA DNA
Importance of Nucleotides
• DNA contains genes, the information needed to synthesize
functional proteins and RNAs
• Building blocks of nucleic acids (RNA, DNA)
• Energy currency in cellular metabolism (ATP: adenosine
triphosphate)
• Allosteric effectors e.g. cAMP
• Structural components of many enzyme cofactors (NAD, FAD)
• Ribosomal RNAs (rRNAs) are components of ribosomes, playing a
role in protein synthesis
• Messenger RNAs (mRNAs) carry genetic information from a gene
to the ribosome
• Transfer RNAs (tRNAs) translate information in mRNA into an
amino acid sequence
• RNAs have other functions, and can in some cases perform
catalysis
ATP is a nucleotide - energy currency
 Structure of nucleotides

Nucleotides have three characteristic components:

[1] A PHOSPHATE GROUP

A nitrogenous base
[2] A PENTOSE SUGAR (pyrimidines or purine)

[3] A NITROGENOUS BASE

A phosphate group

A pentose
sugar
 STRUCTURE OF NUCLEIOSIDE

Remove the phosphate group from nucleiotide, and you have

a nucleoside
Nucleosides are composed of a base (adenine, guanine, cytosine,
thymine, or uracil) attached to a sugar (ribose for ribonucleosides
or deoxyribose for deoxyribonucleosides)

Nucleosides = Sugar + Base (no phosphate)

There two types of nucleic acids

The composition of DNA differs from that of RNA in only two

respects
1. Sugar: DNA containing deoxyribose, while RNA contain ribose
2. Base composition:
DNA: adenine, guanine, cytosine, and thymine
RNA: adenine, guanine, cytosine, and uracil
Pyrimidine and purine
• The bases are abbreviated by their first letters (A, G, C, T, U)
• The purines (A, G) occur in both RNA and DNA
• Among the pyrimidines, C occurs in both RNA and DNA, but T
occurs in DNA, and U occurs in RNA
Some minor bases

• 5-Methylcytidine occurs in DNA of animals and higher plants

• N6-methyladenosine occurs in bacterial DNA
• Bases attach to the C-1 of ribose or deoxyribose
• The pyrimidines attach to the pentose via the N-1 position of the
pyrimidine ring
• The purines attach through the N-9 position
Nucleic Acid
• Nucleotide monomers can link
together via phosphodiester linkage
formed between the 3-OH of
nucleotide and the 5-phosphate of
next nucleotide

• The polynucleotide or nucleic acid backbone thus consists of alternating

phosphate and pentose residues
• The bases are analogous to side chains of amino acids; they vary without
changing the covalent backbone structure
• Sequence is written from the 5' to 3' end: 5'-ATGCTAGC-3'
• Note that the backbone is polyanionic
Interstrand H-bonding between DNA bases
Chemistry of Nucleic Acid
Stability of Nucleic Acids:

➢ H-bonds between the base pairs: Not solely responsible for

overall stability of the helix
➢ Bases can form equally strong and energetically favorable H
bonds with water
➢ dsDNA will only form if the strands are complementary
➢ More than half of the energy is because of base stacking
➢ Nitogenous bases are nonpolar & hence ‘hydrophobic’
➢ Insoluble in aqueous cell environment
➢ It is energetically favorable to exclude water (by stacking the
bases) from the flat surface of bases
➢ Stacking is maximised in dsDNA and Hydrophobic effect
ensures that this is the most energetically favorable arrangement
(Hydrophobic core of stacked bases)
Effect of Acid:
Strong : Hydrolysis to 3 components
Dilute :selective hydrolysis; glycosylic bonds between purine bases
and ribose are broken → Nucleic acids become apurinic
Application: Maxam & Gilbert Sequencing

Effect of Alkali:
DNA
Denaturation
Tautomeric shift (keto→ enol)
RNA
Hydrolysis (2’OH)→ Cyclicphosphodiester: RNA is less stable
because of this.
Spectroscopic and Thermal Properties of DNA
UV Absorption:
• Due to N bases (aromatic)
• Not the sugar-phosphate backbone
• λmax= 260 nm
Application:
• Detection
• Quantity check
• Purity Check
Hypochromicity: (less coloured)
•Absorbance at 260 nm:
Isolated nucleotides > ss DNA / RNA > ds DNA
Base stacking
Maximum intermediate minimum
Hyperchromicity: (more coloured)
•Absorbance at 260 nm:
ds DNA > ss DNA / RNA > Isolated nucleotides
minimum intermediate Maximum
Nucleotides and Nucleic Acids Undergo
Nonenzymatic Transformations
• Alterations in DNA structure that
produce permanent changes in the
genetic information encoded therein are
called mutations
• Evidence suggests an intimate link
between the accumulation of mutations
in an individual organism and the
processes of aging and carcinogenesis
• Several nucleotide bases undergo
spontaneous loss of their exocyclic
amino groups
• Under typical cellular conditions,
deamination of cytosine (in DNA) to
uracil occurs in about one of every 107
cytidine residues in 24 hours, it is the
reason why DNA contains thymine
rather than uracil
• Uracil is readily recognized as foreign in
DNA and is removed by a repair system
 Effect of UV light

• UV light induces the condensation of

two ethylene groups of pyrimidine
bases in nucleic acids forms
cyclobutane pyrimidine dimers
• Formation of a cyclobutane pyrimidine
dimer introduces a bend or kink into
the DNA
• This thymine dimer
interfere in the DNA
replication
• Photolyase is the enzyme
which can break this
dimer
DNA Structure and Types
DNA Structure

Watson and Crick discovered the double helix by

building models to conform to X-ray data

• By the beginnings of the 1950’s, the race was on to move from the
structure of a single DNA strand to the three-dimensional structure
of DNA.
• Among the scientists working on the problem were Linus Pauling,
in California, and Maurice Wilkins and Rosalind Franklin, in
London.
• Watson and crick proposed final structure of DNA on 28 Feb 1953
and received Nobel Prize in 1962
• Maurice Wilkins and Rosalind Franklin used X-ray
crystallography to study the structure of DNA
– In this technique, X-rays are diffracted as they passed through
aligned fibers of purified DNA.
– The diffraction pattern can be used to deduce the three-
dimensional shape of molecules.

• James Watson “learned”

from their research
that DNA was helical
in shape and he deduced
the width of the helix
and the spacing of bases.
Fig. 16.4
• Watson and his colleague Francis Crick began to work on a model
of DNA with two strands, the double helix.
• Using molecular models made of wire, they first tried to place
the sugar-phosphate chains on the inside.
• However, this did not fit the X-ray measurements and other
information on the chemistry of DNA.
• The key breakthrough came when Watson put the sugar-
phosphate chain on the outside and the nitrogen bases on
the inside of the double helix.
– The sugar-phosphate chains of each strand are like the side
ropes of a rope ladder.
– Pairs of nitrogen bases, one from each strand, form rungs.
– The ladder forms a twist every ten bases.
Fig. 16.5
The salient features of DNA double helix

➢Two long polynucleotide chains coiled around a central

axis, forming a right-handed double helix
➢Each poly-deoxyribonucleotide is composed of many
deoxyribonucleotides joined by phosphodiester linkage
between their sugar and phosphate residues
➢Where ever Adenine occurs Thymine is present on the
corresponding position of the other strand and vice-versa
➢Where ever Guanine occurs Cytosine is present on the
corresponding position of the other strand and vice-versa
➢The nitrogenous bases of opposite strands are paired to one
another by hydrogen bonds.
Continue…
• The two chains are antiparallel; that is, each chain has a
specific orientation, and these run in opposite directions.
• The bases of both chains are flat structures, lying
perpendicular to the axis. They are ‘stacked’ on one another,
0.34 nm (3.4 Å) apart, and are located on the inside of the
helix.
• Each complete turn of the helix is 3.4 nm (34 Å) long. This
means that just over ten bases from each strand (10.4 bp)
form one complete turn of the helix.
• Along the molecule, alternating larger major grooves and
smaller minor grooves are apparent.
• The double helix measures approximately 2 nm (20 Å) in
diameter.
Types of DNA
RNA Structure and Types
• In all procaryotic and eucaryotic organisms, 3 general
types of RNAs are found : ribosomal, transfer and
messenger RNAs.
• All 3 types of RNA molecules differ from each other
by size, function and general stability.
 RNA types & functions
Types of RNAs
mRNA - messenger
rRNA - ribosomal
t-RNA - transfer
hnRNA - heterogeneous nuclear
scRNA - small cytoplasmic
snRNA - small nuclear
snoRNA - small nucleolar
regulatory RNAs (siRNA,
miRNA, etc.)
L Samaraweera 2005
Primary Function(s)
translation (protein synthesis) regulatory
translation (protein synthesis) <catalytic>
translation (protein synthesis)
precursors & intermediates of mature mRNAs
& other RNAs
signal recognition particle (SRP)
tRNA processing <catalytic>
mRNA processing, poly A addition <catalytic>
rRNA processing/maturation/methylation
regulation of transcription and translation,
 RNA Structure

• RNA primary structure

• 5’ to 3’ list of covalently linked nucleotides, named by the
attached base
Secondary Structure
• Base are pairing with each other on the basis of complementary
• The double helical secondary structures in RNA can form within
a single RNA molecule or between 2 separate RNA molecules
• Single stranded bases within a stem are called a bulge or bulge
loop if the single stranded bases are on only one side of the stem
• If single stranded bases interrupt both sides of a stem, they are
called an internal (interior) loop.
RNA secondary structure representation
 Ribosomal RNA (rRNA) or Insoluble RNA
• Most stable form of RNA and found in ribosomes
• Ribosomal RNA is most abundant of all types of RNAs and
makes up about 80% of the total RNA of a cell
• Ribosomal RNA represents about 40-60% of the total weight of
ribosomes
• rRNA from all sources has G-C contents more than 50%
 Transfer RNA (tRNA) or Soluble RNA (sRNA)
• The smallest polymeric form of RNA
• tRNA occupy about 15% of the total RNA of the cell
• The most important function of tRNA is to act as specific carriers
of activated amino acids to specific sites on the protein-
synthesizing templates
• Since the code is degenerate, there may also be more than one
tRNA for a specific amino acid
• In a bacterial cell, there are more than 70 tRNAs and in eukaryotic
cells, this number is even greater, because there are tRNAs
specific of mitochondria and chloroplasts
• There are generally several tRNAs specific of the same amino acid
(sometimes up to 4 or 5); they are called isoacceptor tRNAs
• Each tRNA is specific of an amino aicd, i.e., it can bind (or
“accept”) only that particular amino acid, alanyl-tRNAAla or
simply alanyl-tRNA
Common Structural Features
• All tRNA molecules have a common design and consist of 3 folds giving it
a shape of the cloverleaf with four arms; the longer tRNAs have a short
fifth or extra arm. The actual 3-dimensional structure of a tRNA looks
more like a twisted L than a cloverleaf
• All tRNA molecules are unbranched
chains containing from 73 to 93
ribonucleotide residues, corresponding
to M.W. between 24,000 and 31,000
• They contain from 7 to 15 unusual
modified bases. Many of these
unusual bases are methylated or
dimethylated derivatives of A, U, G
and C. These include nucleotides of
pseudouridine, various methylated
adenines and guanines, methylated
pyrimidines such as thymine and 5-
methylcytosine and others
• The 5′ end of tRNAs is phosphorylated. The 5′ terminal residue is usually
guanylate (pG)
• The base sequence at the 3′ end of all tRNAs is CCA. All amino acids bind
to this terminal adenosine via the 3′-OH group of its ribose
• The 4 loops are recognition sites
• A unique similarity among all tRNA molecules is that the overall distance
from CCA at one end to the anticodon at the other end is constant
• About 50% of the nucleotides in tRNAs are base-paired to form double
helices
• However, 5 groups of bases which are not base-paired. These 5 groups, of
which 4 form ‘loops’, are :
1. the 3′ CCA terminal region,
2. the ribothymine-pseudouracil-cytosine ( = T φ C) loop,
3. the ‘extra arm’ or little loop, which contains a variable number of
residues,
4. the dihydrouracil ( = DHU) loop, which contains several dihydrouracil
residues, and
5. the anticodon loop, which consists of 7 bases with the sequence
 Messenger RNA (mRNA) or Template RNA
• Genetic information of nuclear DNA is transmitted to mRNA
which functions as the sites of protein synthesis
• In 1961, the two Nobel laureates, Francois Jacob and Jacques
Monod observed that template RNA do not accumulate as
compared to DNA, rRNA and tRNA, which they called “the
messenger” RNA (mRNA)
• “Messenger” should have the following properties
1. The messenger should be a polynucleotide
2. The base composition of the messenger should reflect the base
composition of the DNA that specifies it
3. The mRNA should be very heterogeneous in size because genes or
groups of genes vary in length. They also correctly assumed that 3
nucleotides code for one amino acid
4. The messenger should be, for a short period, associated with ribosomes
5. The messenger should be synthesized and degraded very rapidly
• All these properties are nowadays ascribed to the messenger RNA
because the other 2 types of RNAs (rRNA and tRNA)are
homogeneous and also their base composition is similar in species
that have very different DNA base ratios
• Messenger RNA is most heterogeneous in size and stability
among all the types of RNAs
• In E. coli, the average size of mRNA is 900 to 1,500 nucleotide
units
• If mRNA carries the codes for the synthesis of simple protein
molecule, it is called monocistronic type and if it codes for more
than one kind of protein, it is known as polycistronic type
• The mRNAs are unstable in the bacterial systems with a half-life
from a few seconds to about 2 minutes.
• In mammalian systems, however, mRNA molecules are more
stable with a half-life ranging from a few hours to one day
 Heterogeneous Nuclear RNA(hnRNA)
• In mammalian cells including those of human beings, a precursor
RNA is first synthesized in the nucleoplasm by DNA-dependent
RNA polymerase
• This precursor is then degraded by a nuclear nuclease to mRNA
that is then translocated to the cytoplasm where it becomes
associated to the ribosomal system
• This precursor RNA constitutes the fourth class of RNA molecules
and is designated as heterogeneous nuclear RNA (hnRNA)
• The hnRNA molecules may have molecular weights exceeding 107
daltons whereas the mRNA molecules are generally smaller than
2×106 daltons
• Most mammalian mRNA molecules are 400−4,000 nucleotides in
length whereas a hnRNA molecule possesses 5,000−50,000
nucleotides
Types of secondary and tertiary
structures of nucleotides
• Certain DNA Sequences Adopt Unusual Structures
• A sequence of alternating T and C residues can be considered a mirror repeat
centered about a central T or C
• These sequences form an unusual structure in which the strands in one half of the
mirror repeat are separated and the pyrimidine containing strand (alternating T
and C residues) folds back on the other half of the repeat to form a triple helix
• The purine strand (alternating A and G residues) is left unpaired. This structure
produces a sharp bend in the DNA