DNA - Structure

A K A Mandal

A K A Mandal 1
 What is DNA?

• DNA is a Nucleic Acid

• Polymer of Nucleotides
• Each nucleotide consists of
– Deoxyribose (5-carbon sugar)
– Phosphate group
– A nitrogen-containing base
• Four bases
– Adenine, Guanine, Thymine, Cytosine
A K A Mandal 2
 Nucleic Acids
• A nucleic acid is a polynucleotide – a polymer of
nucleotides
• Each nucleotide has three components-
• A cyclic five-carbon sugar-
– ribose in RNA, deoxyribose in DNA [ribose has 2’-
OH and deoxyribose has 2’-H, this difference
makes DNA chemically more stable than RNA]
• A purine or pyrimidine base-
– Attached to the 1’-carbon atom of the sugar by an
N-glycosidic bond. Adenin (A) and guanin (G) are
purine base and cytosine (C), thymine (T), and
uracil (U) are pyrimidine base. A, G and C are
present in both DNA and RNA. T is found only in
DNA and U is only in RNA. Exception: in some t-
RNA T is present while, few phage DNA contain U.
Nucleotide Structure
• A phosphate-
– Attached to 5’ carbon of sugar by a phosphoester
linkage. This phosphate is responsible for the
strong negative charge of nucleotides and nucleic
acids
Nucleoside = Pentose sugar + N-base
A K A Mandal Nucleotide = Nucleoside + phosphates 3
 Structure of DNA/RNA - Bases and Sugars

pyrimidines

C U T

purines

G A

Ribose
sugars
A K A Mandal 4
 The polymerization reaction that results
in synthesis of a DNA polynucleotide
• The nucelotides in nucleic acids are
covalently linked by a second
phosphoester bond that joins the 5’-
phosphate of one nucleotode and the 3’-
OH group of the adjacent nucleotide.
Thus the phosphate is esterified to both
the 3’- and 5’-carbon atoms; this unit is
often called a phosphodiester group
• The purine and pyrimidine bases are not
engaged in any covalent bond with
each other. Thus a polynucleotide
consists of alternating sugar-phosphate A short DNA polynucleotide
backbone having 3’-OH terminus and showing the structure of the
one 5’-phosphate terminus phosphodiester bond.
Note that the two ends of the
A K A Mandal polynucleotide are chemically
5
distinct
 The polymerization reaction that results in
synthesis of a DNA polynucleotide
• A nucleoside consists of a purine or pyrimidine base linked to
position 1 of a pentose sugar.
• Prime (‘) to distinguish them
• The difference between DNA ad RNA is in the group at the 2’
position of sugar. DNA has a deoxyribose sugar (2’-H); RNA has a
ribose sugar (2’-OH)
• A nucleotide consists of a nucleoside linked to a phosphate group
on either the 5’ or 3’ position of the (deoxy)ribose
• Successive (deoxy)ribose residues of a polynucleotide chain are
joined by a phosphate group between the 3’ position of one sugar
and the 5’ position of the next sugar
• One end of the chain (conventionally left) has a free 5’ end and the
other has free 3’ end
• DNA contains four bases adenine, guanine, cytosine, and thymine;
RNA has uracil instead of thymine
A K A Mandal 6
 The polymerization reaction that results in
synthesis of a DNA polynucleotide
•Synthesis occurs in the 5′→3′
direction, new nucleotide being
added to the 3′-carbon at the end
of the existing polynucleotide. β-
and γ-phosphates of the
nucleotide are removed as a
pyrophosphate molecule.

•Polymerization reaction involves

removal of the two outer
phosphates (the β- and γ-
phosphates) from one nucleotide
and replacement of the hydroxyl
group attached to the 3′-carbon of
theA Ksecond
A Mandal nucleotide 7
3’ to 5’ Phosphodiester Bonds Make the
Sugar-Phosphate Backbone

New monomers
add here

Strand has 5’-PO4

end and 3’-OH end
A K A Mandal 8
• Phosphodiester Bonds
– Phosphate is from phosphoric acid
– Hydroxyl groups on sugars represent alcohol
• Acid and alcohol gives the ester
– Phosphate reacts with two –OH groups
– One water molecule is removed

A K A Mandal 9
 The polymerization reaction that results in synthesis of a
DNA polynucleotide

• Two ends of the polynucleotide are chemically distinct:

– 1)unreacted triphosphate group at 5′-carbon (5′ or 5′-P terminus)
– 2)unreacted hydroxyl at 3′-carbon (3′ or 3′-OH terminus).
This means that the polynucleotide has a chemical direction,
expressed as either 5′→3′ or 3′→5′

• An important consequence of the polarity of the

phosphodiester bond is that the chemical reaction needed to
extend a DNA polymer in the 5′→3′ direction is different to
that needed to make a 3′→5′ extension.

All natural DNA polymerase enzymes are only able to carry out
5′→3′ synthesis, which adds significant complications to the
process by which double-stranded DNA is replicated. The
same limitation applies to RNA polymerases, the enzymes
which make RNA copies of DNA molecules
A K A Mandal 10
 Nucleic Acid Nomenclature
Purines

Base Nucleoside Nucleotide

Adenine (A) Adenosine (rA) Adenylic acid or

adenosine
monophosphate (AMP)
Deoxyadenosine (dA) Deoxyadenylic acid or
deoxyadenosine
monophosphate (dAMP)
Guanine (G) Guanosine (rG) Guanylic acid or
guanosine
monophosphate (GMP)
Deoxyguanosine (dG) Deoxyguanylic acid or
deoxyguanosine
monophosphate (dGMP)
A K A Mandal 11
 Nucleic Acid Nomenclature
Pyrimidines

Base Nucleoside Nucleotide

Cytosine (C) Cytidine (rC) Cytidylic acid or Cytidine

monophosphate (CMP)

Deoxycytidine (dC) Deoxycytidylic acid or

deoxycytidine
monophosphate (dCMP)
Thymine (T) Thymidine(dT) Thymidylic acid or
thymidine
monophosphate (TMP)
Uracil (U) Uridine (rU) Uridylic acid or uridine
monophosphate (UMP)

A K A Mandal 12
Biomolecules - Nucleic Acid - Polymerization of nucleotides

• https://www.youtube.com/watch?v=0SErT
E3vpRg

A K A Mandal 13
 DNA Double Helix

A K A Mandal 14
 DNA – Structure?

• The Race is On
– Linus Pauling
– Maurice Wilkins and Rosalind Franklin
– Watson and Crick

A K A Mandal 15
 Linus Pauling and
Robert Corey at
Cal Tech were in
the Race to
Determine DNA
structure

A K A Mandal 16
 Rosalind Franklin

A K A Mandal 17
 James Watson and Francis Crick Used
Molecular Modeling to Deduce DNA
structure

A K A Mandal 18
 DNA Double Helix - History
• Before 1950, various lines of evidence had
shown that cellular DNA molecules are
comprised of two or more polynucleotides
assembled together in some way
• Linus Pauling proposed an incorrect triple helix
model
• Finally, double helix, discovered by Watson and
Crick on Saturday 7 March 1953
The single most important breakthrough in
biology during the 20th century
A K A Mandal 19
 The evidence that led to the double helix

• Watson and Crick used four types of information to

deduce the double helix structure
– Biophysical data of various kinds. The water content of DNA
fibers was particularly important because it enabled the density of
the DNA in a fiber to be estimated
– X-ray diffraction patterns most of which were produced by
Rosalind Franklin of Kings College, London revealed the helical
nature of the structure and indicated some of the key dimensions
within the helix
– The base ratios, discovered by Erwin Chargaff of Columbia
University, New York, showed that, although the values are
different in different organisms, the amount of A is always the
same as the amount of T, and the amount of G equals the amount
of C
– Model building, which was the only major technique that Watson
and Crick made use of themselves 20
 Composition of DNA
• Chargaff showed:
– Amount of adenine relative to guanine differs among species
– Amount of adenine always equals amount of thymine and
amount of guanine always equals amount of cytosine
– Results
A = T, C = G
A + G = C + T (purine = pyrimidine)
A + T does not equal C + G {ie (A=T) ≠ (G=C)}

– Members of a species similar but different species vary in

AT/CG ratio
Chargaff’s Base Pairing Rule

A K A Mandal 21
 X ray diffraction
• When X rays are focused through isolated macromolecules or
crystals of purified molecules, the X rays are deflected by the atom
of the molecules in specific patterns called diffraction patterns.

• It provides the information about the organization of the components

of the molecules.

• Watson and Crick had X ray crystallographic data on DNA structure

from the studies of Wilkins and Franklin and their coworkers.

• These data indicated that DNA was a highly ordered, multiple

stranded structure with repeating sub structures spaced every 3.4 Å
(1 Angstrom = 10-8 cm )

A K A Mandal 22
 Rosalind Franklin’s Work
• Was an expert in X-ray crystallography
• Used this technique to examine DNA fibers
• Concluded that DNA was some sort of helix
• Rosalind Franklin’s data provide clues about
DNA’s 3-D shape
– Helix
– Width = 2 nm  probably two strands (DOUBLE
HELIX)
– Nitrogenous bases = 0.34 nM apart
– One turn every 3.4 nM (10 base pairs per turn)

One of the X-ray diffraction photographs of DNA that

lead to the double helix model of DNA structure. An
X-ray crystallographer can recognize the central
cross-shaped pattern as indicative of helical
structure. The heavy dark patterns (top and bottom)
Indicate that the bases are stacked perpendicular to
the axis of the molecule with a periodicity of 3.4Å.
A K A Mandal 23
• The arrangement of the three major
components in nucleic acid polymers was
already well known – but the 3-D shape
was still unclear
– Sugar phosphate backbone
– Bases

A K A Mandal 24
• Putting the hydrophobic nitrogenous bases
on the inside, and the sugar-phosphate
groups on the outside was a stable
arrangement

A K A Mandal 25
• Base pairing was worked out by trial
and error
– The distance between the sugar-
phosphate backbone groups is constant
• Therefore purine-purine or pyrimidine-
pyrimidine were not allowed because
spacing would be in inconsistent with data
– Purines = A and G (two organic rings)
– Pyrimidines – C and T ( one organic ring)

A K A Mandal 26
 • Purine-pyrimidine base pairing would be
consistent with X-ray data

A K A Mandal 27
 • Hydrogen bonding between purines and
pyrimidines established the appropriate pairs and
reinforced Chargaff’s Rules
– 2 hydrogen bonds between
A and T
– 3 hydrogen bonds between
G and C

A K A Mandal 28
 DNA is a Double Helix
Three notions converged in the construction of the double helix
model for DNA by Watson & Crick (1953):

• 1. X-ray diffraction data showed that DNA has the form of a regular
helix, making a complete turn every 34 Å (3.4 nm) with a diameter of
~20 Å (2 nm). Since the distance between adjacent nucleotides is
3.4 Å, there must be 10 nucleotides per turn
• 2. The density of DNA suggests that the helix must contain two
polynucleotide chains. The constant diameter of the helix can be
explained if the bases in each chain face inward and are restricted
so that a purine is always opposite to a pyrimidine, avoiding
partnerships of purine-purine (too wide) or pyrimidine- pyrimidine
(too narrow)
• 3. Irrespective of the absolute amounts of each base, the proportion
of G is always equal to C in DNA and A is always equal to T.

A K A Mandal 29
 Nature 171: 737-738 – April
1953
• Watson JD and
Crick FC (1953)
Molecular
Structure of
Nucleic Acids: A
Structure for
Deoxyribose
Nucleic Acid.
• 1962 – Nobel
Prize awarded
to three men –
Watson, Crick
and Wilkins 30
 Watson-Crick Model - Double Helix:
Key Features
• DNA consists of two nucleotide
strands: Double Helix
• Strands run in opposite directions –
Antiparallel. One strand runs in the 5’
to 3’ direction, whereas its partner runs
3’ to 5’
• Strands are held together by hydrogen
bonds between bases
• A binds with T and C with G
• The sugar-phosphate backbone is on
the outside and carries negative
charges on the phosphate groups.
[when DNA is in solution in vitro, the
charges are neutralized by the binding
of metal ions, typically by Na+. In the
cell, positively charged proteins
provide some of the neutralizing force.]
A K A Mandal 31
 Watson-Crick Model - Double Helix: Key Features

• The bases lie on inside. They are flat

structures, lying in pairs
perpendicular to the axis of the helix.
Proceeding along the helix, bases are
stacked above one another like a pile
of plates
• The high degree of stability of DNA
double helices results in part from the
large number of hydrogen bonds
between the base-pairs and in part
from the hydrophobic bonding (or
“stacking forces”) between the
stacked base-pairs.

A K A Mandal 32
Watson-Crick Model - Double Helix: Key Features

• Each base pair is rotated ~36o

around the axis of the helix relative to
the next base pair. So 10 base pairs
make a complete turn of 360o
• The twisting of the two strands
around one another forms a double
helix with a minor groove (~12 Å )
and a major groove (~22 Å across).
The double helix is a right handed;
the turns are clockwise looking along
the helix axis
• These features represent the
accepted model for what is known as
the B form of DNA
33
 DNA is antiparallel

A K A Mandal 34
 Base-pairing rule
The four bases of DNA are:
Adenine (A) Guanine (G) Thymine
(T) Cytosine (C)
Adenine always hydrogen bonds
with Thymine (A-T)
Guanine always hydrogen bonds
with Cytosine (G-C)
These bonding patterns are called
base pairings (bp) and the paired
bases (G with C, or A with T) are said
to be complementary
A K A Mandal 35
 The double helix has structural flexibility

• The double helix described by Watson

and Crick is called the B-form of DNA.
The DNA in living cells is thought to be
predominantly in this B-form
• It is now clear that genomic DNA
molecules are not entirely uniform in
structure mainly because each nucleotide
in the helix has the flexibility to take up
slightly different molecular shapes:
– rotation around the β-N-glycosidic bond
– changing the orientation of the base relative to
the sugar, and
– rotation around the bond between the 3′- and
Computer-generated images
4′-carbons of B-DNA (left), A-DNA (center),
Z-DNA (right)
A K A Mandal 36
A K A Mandal 37
 The double helix has structural
flexibility
• In high concentration of salts or in a
dehydrated state, (75% humidity) DNA
exists in the A- form
• Recently, certain DNA sequences have
been shown to exist in a unique left
handed, double helical form called
Z-DNA.
• The helices of A and B form DNA are
wound in a right handed manner
A K A Mandal 38
 Features of different conformations of the DNA
double helix

Feature B-DNA A-DNA Z-DNA

Type of helix Right-handed Right-handed Left-handed

Helical diameter (nm) 2.37 2.55 1.84

Rise per base pair (nm) 0.34 0.29 0.37

Distance per complete turn (pitch)

3.4 3.2 4.5
(nm)
Number of base pairs per complete
10 11 12
turn

Topology of major groove Wide, deep Narrow, deep Flat

Topology of minor groove Narrow,shallow Broad, shallow Narrow,deep

A K A Mandal 39
 A form of DNA

• 11 bp / rotation
• Axis tilted considerably 20.2 °
• Hence displacement of groove depth is increased and shallow
groove decreased
• Axial raise 2.56 A
• A form is a less hydrated form of B form
• Shorter, wider helix than B.
• Deep narrow major groove not easily accessible to proteins
• Wide, shallow minor groove accessible to proteins, but lower
information content than major groove.
• Favored conformation at low water concentrations
• Base pairs tilted to helix axis

A K A Mandal 40
 B Form of DNA
• B DNA – proposed by Watson and Crick Model of DNA
• It is a native conformation in a solution
• X ray Difraction structure is obtained by 66% of Humidity
• Right handed and anti parrelle double helix structure with sugar
phosphate backbone
• Purine pyramidins structure is roughly perpendicular to the
backbone
• Tilt of bp is normal 6.3°
• 10 bp / rotation
• Axial raise is 3.37 A
• Most common DNA conformation in vivo
• Narrower, more elongated helix than A.
• Wide major groove easily accessible to proteins
• Narrow minor groove
• Favored conformation at high water concentrations
• Base pairs nearly perpendicular to helix axis
A K A Mandal 41
 Z DNA
• Alexander Rich and his colleagues discovered in 1979 that
DNA does not always have to be right-handed. They showed
that double stranded DNA containing strands of alternating
purines and pyrimidines (e.g., poly[dG-dC] . poly[dG-dC]):
—GCGCGCGC —
—CGCGCGCG —
can exist in an extended left-handed helical form.
• Although Rich discovered Z-DNA in studies of model
compounds like poly[dG-dC] . poly[dG-dC], this structure
seems to be more than just a laboratory curiosity.
• Evidence suggests that living cells contain a small proportion
of Z-DNA.

A K A Mandal 42
 Z form of DNA
• Z DNA helix is left handed and has a structure that
repeats every 2 base pairs.
• The major and minor grooves show little difference in
width.
• Formation of this structure is generally unfavourable,
although conditions promote it;
• The Z-DNA conformation has been difficult to study
because it does not exist as a stable feature of the
double helix. Instead, it is a transient structure that is
occasionally induced by biological activity and then
quickly disappears
• The Z-DNA has been found in a large number of living
organisms including mammals, protozoans and several
plant species.
A K A Mandal 43
 C form of DNA

• Reduction of hydration of B form below

66% results in C form
• Helix symmetry (number of base pairs per
turn) is 28/3 i.e. 9 and 1/3 bp/rotation
• Axial raise 3.32 Å
• Tilt (negative) is 7.8°

A K A Mandal 44
 D form of DNA

• Form Poly dT and Poly G(dC and dG) and

8bp/rotation
• As in C forms base pairs displaced to axis
helix backwards relative the of the helix.
Axis tilted considerably 16.7°
• Axial raise 3.03 Å

A K A Mandal 45
The pattern of base pairing is the mechanism by which DNA
holds information.
Humans have a > 6 billion of these base pairings
Less than 5% of our DNA actually forms genes
There about 30,000 genes encoded in our DNA, nearly half
of these genes either have yet to be discovered or their
function is unknown

A K A Mandal 46
 Reading DNA Strands
Single strand of DNA:
5’-AGCATTCG-3’

3’-TCGTAAGC-5’
Complementary strand of above, usually written 5’ to 3’:
5’-CGAATGCT-3’

Double-stranded fragment is written:

5’-AGCATTCG-3’
3’-TCGTAAGC-5’
A K A Mandal 47
 Circular DNA
• Some DNAs are circular:
– small monkey DNA virus SV40 (circular, double-
helical DNA mol of 5,000 bp),
– Most bacterial chromosomes
– Many bacteria have small, autonomously replicationg
genetic elements known as plasmids, which are
generally circular DNA molecules
• Some DNA mol are both linear and circular:
bacteriophage lambda, a DNA virus of E. coli
(Lin DS in virion and circular when genome is
injected into E. coli during infection)
A K A Mandal 48
The Structure of DNA: https://www.youtube.com/watch?v=o_-6JXLYS-k

A K A Mandal 49
 References
• Text Books
– 1. Nelson,D.N. and Michal M.C. 2007. Lehninger. Principles of
Biochemistry. W.H. Freeman publications.
– 2. Gardner, E.J., Simmons, M.J., and Snustad. D.P. 2005.
Principles of genetics. 8th edition. Wiley India, Nice Printing
press, New Delhi
– 3. James D. Watson et al.,2004. Molecular biology of the gene.
Pearson education Inc.
• References
– 1. Benjamin Lewin. 2004. Genes VIII. Pearson Education, Inc.
– 2. Geoffrey M. Cooper. 2000. The Cell: A Molecular Approach.
2nd Edition. Sinauer Associates, Inc.
– 3. Lodish H. et. al., 2004. Molecular Cell Biology, 5th Edition,
W.H. Freeman & Co.
A K A Mandal 50