BCH 202

GENERAL BIOCHEMISTRY II
CHEMISTRY OF NUCLEIC ACIDS
 DEFINATION and Structure of Nucleic Acids
• Nucleic acid describes a class of biological molecules
that are polymers; made up of monomeric units
(repeated units) called nucleotides.
STRUCTURE
Nucleotides
• A single Nucleotide is composed of three basic
components:
1. Pentose sugar
2. Nitrogeneous bases
3. Phosphate group
 PENTOSE SUGAR
• The sugar molecule found in nucleic acid is a monosaccharide sugar
that contain five carbon atoms (pentose).
• The two important types of pentoses found in nucleic acids are ribose
(β-D-ribofuranose) and 2-deoxyribose (β-D-2-deoxyribofuranose).

• The important functional groups that differentiate between the two

types of pentose sugars are H and OH groups on carbon 2.
• In ribose, OH group is attached to C2 while deoxyribose lacks an oxygen
atom on C2. The prefix ‘deoxy’ means removal of oxygen.
• DNA contains a deoxyribose sugar thus called DeoxyriboNucleic Acid
while RNA contains ribose pentose sugar hence called RiboNucleic Acid.
 NITROGENOUS BASES
• Human body contains organic compounds
such as Carbon, Hydrogen and Oxygen in
varying amounts.
• Nitrogen combines with these compounds to
form heterocyclic ring structures called
nitrogenous bases.
• Nitrogenous bases are classified into two
categories; purines and pyrimidines.
 PURINES
• Purines have a 6-member heterocyclic
aromatic ring fused to a 5 member imidazole
ring.
• The two major purines types are adenine (A)
and guanine (G).
• Both DNA and RNA contains A and G.
• The substituent at the "top" of the 6-member
ring of the 2-ring system (i.e. at C6) are major
determinants of the H-bonding (or base
pairing) capacity of the purines.
• Adenine is referred to as the "amino" base for
purines, which is 6-aminopurine. The amino
group serves as the H-bond donor during base
pairing with the C2 keto group of thymine or
uracil.
• "keto" base for purine is guanine; note the
keto group at C6.
• The C2 of guanine is bonded to two nitrogen
atoms within the ring and also to an exocyclic
amino group.
• Thus atoms 1, 2, and 3 of guanine form a
guanidino group: The "guan" part of the name
of the guanidino group and of guanine comes
from guano, or bat droppings.
• These excretions are rich sources of purines.
• The N-9 atom of purine base is the site for
attachment to the pentose sugar.
• Purines undergo keto-enol tautomerization,
and the keto tautomer is more prevalent in
nucleic acids.
• The keto-enol tautomerization contributes to
mutations: the enol form will make different
base pairs than the keto form.
 PYRIMIDINES

• Pyrimidines are 6 membered heterocyclic

aromatic ring.
• Two nitrogen atoms are connected to 4 carbon
atoms by a conjugated double bonds, thus
giving the base substantial aromatic character.
• The common pyrimidines found in nucleic acids
are cytosine (C), Thymine (T) and Uracil (U).
• C and T are found in DNA while U is found only
in RNA.
• All the common pyrimidines in DNA and RNA
have a keto group at C2, but they differ in the
substituent at C4, at the "top" of the ring.
• the substituent at C4, as well as N3 of the ring, are
involved in H-bonding to complementary bases in the
secondary structures of nucleic acids.
• Cytosine is referred to as the "amino" pyrimidine base,
because of its exocyclic amino group at C4.
• The "keto" bases are uracil and thymine, again named
because of their keto groups at the top of the ring.
• Thymine is 5-methyl uracil; it is found only in DNA.
• Thymine and uracil are identical at the N3 and C4
positions, and they will both form H-bonds with
adenine.
• N-1 is involved in bond formation with the ribose sugar.
• Pyrimidines can exist in either keto (lactam) or
enol (lactim) tautomer; they exist in the keto
form in nucleic acids.
 NUCLEOSIDES AND NUCLEOTIDES
• Nucleosides have 2 parts: purine or pyrimidine bases attached to
a pentose sugar.
• A nucleotide has 3 parts: a nitrogeneous base attached to a
pentose sugar, attached to a phosphate group; it is a nucleoside
esterified to a phosphate.
• The phosphate is attached by an ester linkage to a hydroxyl group
on the sugar, usually to the 5' or 3' OH.
Note that the atoms in the (deoxy)ribose ring are numbered 1', 2',
3', etc. when in nucleotides or nucleic acids to avoid confusion
with the numbering system of the bases.
• Sometimes the connection with phosphate is at the 2' position in
RNA.
• 1, 2 or 3 phosphates (or more) can be attached to 5' or 3'
position. Starting at the 5'-OH, these phosphates are called α, β, γ.
 NOMENCLATURE OF NUCLEOSIDES AND
NUCLEOTIDES
• The nomenclature for the five types of bases,
nucleosides and nucleotides is as follows:

Purines ‘osine’ pyrimidines ‘dine’

 Nucleoside Conformation
• Rotation about the C-1‘ bond allows two orientations:
• anti- extends the base and pentose rings in opposite directions.
• For pyrimidines, this means that O-2 faces away from the pentose.
• syn- orients the base and the pentose in the same direction.
• Free purine nucleosides, in particular guanosine, favour the syn-
orientation, but adopt the common anti- orientation within most DNA and
RNA helices.
• Pyrimidines adopt anti orientation almost exclusively.
 Polynucleotides
Phosphodiester linkages
• The 3' OH of the (deoxy) ribose of one nucleotide can link to
the 5' OH of the (deoxy) ribose of another nucleotide via a
phosphate ester bond.
• In synthesizing a nucleic acid polymer from individual
monomers, the 3'-hydroxyl group of one nucleotide attacks
the α-phosphate of another nucleotide.
• therefore, a polymer is made by linking nucleotides via
phosphodiester bonds.
• This forms the basis of sugar phosphate backbone of nucleic
acids.
• The sugar phosphate backbone has an orientation that is
denoted by the orientation of the sugars. the chain of
nucleotides runs in a 5' to 3' orientation from left to right.
 CLASSIFICATION AND FUNCTION
• There are two major types of nucleic acids in the
body:
– deoxyribonucleic acid (DNA): found mostly in the
Nucleus of the cell
– ribonucleic acid (RNA): mostly found outside the nucleus
• The two important functions of nucleic acids in the
cell.
1. To store genetic information of the cell necessary
for carrying out genetic functions
2. To transfer genetic information and synthesize
proteins
 DNA
• DNA is double stranded
• It has two polynucleotide strands
wound together, are complementary
and run antiparallel to each other
• i.e One strand run in 5’-3’ direction the
complementary strand run 3’-5’
direction
• The Adenines on one strand form base
pair with Thymines and the Guanines
form base pair with Cytosines of the
other strand
• The permitted hydrogen bonding is:
adenine with thymine (2 hydrogen
bonds); cytosine with guanine (3
hydrogen bonds).
• The model is called Watson and Crick
double helix
Types of DNA double helix
 RNA
• RNA is a single stranded polynucleotide.
• RNA fold back on itself to assume three
dimensional shapes such as Hairpin, clover
leaf etc
• RNA is mostly an adaptor molecule, used to
copy information in DNA (transcription) and
use the information for protein synthesis
(translation) machinery/structure.
 TYPES OF RNA

• Messenger RNA (mRNA) – IMPORTANT for

transcription of the information in the DNA. Used
to carry the information in temporary format from
the DNA archive to the protein synthetic
machinery.
• Ribosomal RNA (rRNA) - This RNA forms the core
of the ribosome, which is used to make proteins.
The ribosome consists of RNA molecules and
many proteins (~100 in eukaryotes).
• Transfer RNA (tRNA) - It bring amino acids into the
ribosome for protein synthesis.
Comparison of DNA and RNA
 Stability of DNA double helix
Forces that stabilize the DNA structure
1) Electrostatic
2) Hydrophobic "forces"
3) H-bonds
4) Stacking interactions
• base pairs are found in the interior of the helix.
• charged and hydrophilic sugar-phosphate backbone
are on the exterior.
• phosphates from the 2 strands are also kept away
from each other (as much as possible) to reduce the
electrostatic repulsion.
 DNA: Effect of temperature
• The two polynucleotide chains of double-helical DNA can
be separated under certain conditions from dsDNA to
ssDNA.
• Terms that describe the change from dsDNA to ssDNA are:
Melting
Denaturation
Strand separation.
• Terms that describe the change from ssDNA to dsDNA are
Annealing
Renaturation
Hybridization.
 DNA DENATURATION
• DNA can be denatured under extreme conditions of temperature
or pH.
• These conditions disrupt the hydrogen bonding and hydrophobic
interactions between the bases, and result in separation of the
strands.
• Denaturation is also referred to as melting, since this transition can
be caused by heating.
 Melting temperature (Tm)
• The melting temperature is defined as the temperature at the midpoint
in denaturation curve (point of separation).
Factors that Influences on Tm:
1. GC content: the higher the GC%, the higher the Tm (G-C base pairs have
3 H-bonds and are thus stronger than A-T base pairs)
2. Salt: the higher the [salt], the higher the Tm (ions shield charges and
thus lessen repulsion between phosphates)
3. Low (<2.3) or high (>11.5) pH decrease Tm (ionization of the bases)
4. ionic strength (ì): The Tm increases as the cation concentration
increases. The phosphodiester backbone’s negative charges tend to
repel each other, but that repulsion is greatly decreased when each
phosphate is surrounded by a cloud of small cations.
5. Organic compounds such as formide or urea, will decrease the Tm by
destabilizing the double helix by competing as H-bond partners or by
disrupting the water shell around the bases
 DNA RENATURATION (annealing)
• Under certain conditions, DNA can be renatured — the
complementary strands are brought back together and the
correct H-bonding pattern is reproduced. This occurs
efficiently at the annealing temperature
• Renaturation is also referred to as annealing; this is favored by
cooling to about 20 to 25°C below the melting temperature
• Annealing temperature = Tm - 25°C
 HYDROLYSIS BY ACID AND BASE

• DNA is generally stable - resists attack by acid and alkali solutions

• In mild acid solutions - at pH 4 - the β-glycosidic bonds of the
purine bases are hydrolyzed; protonation of purine bases (N7 of
guanine, N3 of adenine) occurs at this pH.
• Protonated purines are good leaving groups.
• depurinated sugar can easily isomerize into the open-chain form
and in this form the depurinated (or apurinic) DNA is susceptible
to cleavage by hydroxyl ions.
• DNA is not susceptible to alkaline hydrolysis.
• RNA is very unstable in alkali solutions due to hydrolysis of the
phosphodiester backbone.
• The 2'OH group in ribonucleotides renders RNA molecules
susceptible to strand cleavage in alkali solutions.
 Enzymatic hydrolysis of nucleic acids
• enzymes that hydrolyze nucleic acids are
called nucleases.
• Organs that provide digestive fluids such as
pancreas are rich in nucleases.
• Fungi and snake venom are good sources of a
class of nucleases called phosphodiesterases
• There are many enzymes that cleave RNA –
ribonucleases (Rnases), those that act on DNA
are deoxynucleases; Dnase.
 Flow of genetic information
• DNA→RNA→PROTEINS
• Central dogma of biology