DETAILED STRUCTURE OF DNA

2. DNA
A molecule that contains
the instructions an
organism needs to develop,
live and reproduce.
These instructions are
found inside every cell and
are passed down from their
parents to their children.
Nearly every cell in the
persons body has the same
DNA

3. Made up of molecules called
nucleotides.
Each molecule contains a
phosphate group , a sugar group
and a nitrogen base.
These nitrogen bases are
adenine(A),guanine(G),thymine
(T)and cytosine(C).
The order of these bases is what
determines the DNA’s instructions
or genetic codes.
COMPOSITION OF DNA

4. NUCLEIC ACIDS
 Nucleic acids are
polymers
 Monomer-----nucleotides
Components:-
 Nitrogenous bases
 Purines
Pyrimidines
 Sugars
Ribose
Deoxyribose
 Phosphates
+ nucleosides=nucleotides

5. NUCLEIC ACIDS
 Nucleic Acids, are naturally occurring chemical compound that is
capable of being broken down to yield phosphoric acid, sugars, and
a mixture of organic bases (purines and pyrimidines).
 Nucleic acids are the main information-carrying molecules of the
cell, and, by directing the process of protein synthesis, they
determine the inherited characteristics of every living thing.
 The two main classes of nucleic acids are deoxyribonucleic acid
(DNA) and ribonucleic acid (RNA).
 DNA is the master blueprint for life and constitutes the genetic
material in all free-living organisms and most viruses.
 RNA is the genetic material of certain viruses, but it is also found
in all living cells, where it plays an important role in certain
processes such as the making of proteins.

6.  Nucleic acids are polynucleotide—that is, long chainlike molecules composed
of a series of nearly identical building blocks called nucleotides.
 Each nucleotide consists of a nitrogen-containing aromatic base attached to a
pentose (five-carbon) sugar, which is in turn attached to a phosphate group.
Each nucleic acid contains four of five possible nitrogen
containing bases: adenine (A), guanine (G), cytosine (C),
thymine (T)and Uracil(U).
 A and G are categorized as purines, and C, T, and U are collectively
called pyrimidines. All nucleic acids contain the bases A, C, and G; T, however,
T is found only in DNA, while U is found in RNA.
 The pentose sugar in DNA (2′-deoxyribose) differs from the sugar in RNA
(ribose) by the absence of a hydroxyl group (−OH) on the 2′ carbon of the
sugar ring. Without an attached phosphate group, the sugar attached to one
of the bases is known as a nucleoside.
 The phosphate group connects successive sugar residues by bridging the 5′-
hydroxyl group on one sugar to the 3′-hydroxyl group of the next sugar in
the chain. These nucleoside linkages are called phosphodiester bonds and are
the same in RNA and DNA.

7. THE SUGARS
(in DNA)
(in RNA)
 DEOXYRIBOSE
 Deoxyribose is a pentose sugar of RNA with five sugars . Four out
of five carbon atoms plus an oxygen atom forms a five membered
ring
 The fifth carbon is outside the group and forms the part of the –
CH2 group.
 Deoxyribose has only two (-OH) groups (on carbons 3’ and 5’) and
thus can only form two deoxyribonucleotides , the 3’ and 5’
phosphate derivatives.
 RIBOSE
 Ribose is a pentose sugar of RNA with five carbons.
 It has an identical structure to DNA except there is a (-OH) group
instead of hydrogen on carbon atom 2’.
 Ribose has free (-OH) groups on carbons 2’, 3’ and 5’.
 The phosphate can attach to any of these three positions.

8. THE BASES
 The main nitrogenous bases present in DNA and RNA are purines
and pyrimidines.
 PURINES
 A series of heterocyclic compounds that are variously substituted in
nature are known also as purine bases .
 They include adenine and guanine as constituents of nucleic acids
and many alkaloids.
PYRIMIDINES
 A heterocyclic compound C4H4N2,that is the basis of several
important biochemical substances.
 They include cytosine , thymine and uracil as constituents of nucleic
acids.

9. PURINES AND PYRIMIDINES
PURINES PYRIMIDINES

10. TAUTOMERIC FORMS OF BASES:SOME OF THE POSSIBLE
TAUTOMERIC FORMS OF (a)guanine and (b)thymine. Cytosine and
adenine can also undergo similar proton shifts.
Guanine (enol or lactim form)=Guanine (keto or lactam form)
Thymine(enol or lactim form)=Thymine (keto or lactam form).

13. NUCLEOTIDES-THE BUILDING BLOCKS OF DNA
NUCLEOTIDES, any member of a class of organic
compounds in which the molecular structure
comprises a nitrogen-containing unit (base)
linked to a sugar and a phosphate group.
 The nucleotides are of great importance to living
organisms, as they are the building blocks of
nucleic acids, the substances that control all
hereditary characteristics.
 Several nucleotides are coenzymes; they act with
enzymes to speed up (catalyze) biochemical
reactions.

14.  Nucleotides are synthesized from readily available precursors in the cell.
 The ribose phosphate portion of both purine and pyrimidine nucleotides is
synthesized from glucose via the pentose phosphate pathway.
 The six-atom pyrimidine ring is synthesized first and subsequently attached to
the ribose phosphate.
 The two rings in purines are synthesized while attached to the ribose phosphate
during the assembly of adenine or guanine nucleosides.
 In both cases the end product is a nucleoside carrying a phosphate attached to
the 5′ carbon on the sugar. Finally, a specialized enzyme called a kinase adds two
phosphate groups using adenosine triphosphate (ATP) as the phosphate donor to
form ribonucleoside triphosphate, the immediate precursor of RNA. For DNA,
the 2′-hydroxyl group is removed from the ribonucleoside diphosphate to give
deoxyribonucleoside diphosphate. An additional phosphate group from ATP is
then added by another kinase to form a deoxyribonucleoside triphosphate, the
immediate precursor of DNA.
 During normal cell metabolism, RNA is constantly being made and broken down.
The purine and pyrimidine residues are reused by several salvage pathways to
make more genetic material. Purine is salvaged in the form of the corresponding
nucleotide, whereas pyrimidine is salvaged as the nucleoside.

15. NUCLEOTIDES AND NUCLEOSIDES

17. THE SEVEN TORSION ANGLES THAT DETERMINE THE
CONFORMATION OF A NUCLEOTIDE UNIT
The conformation of a nucleotide
unit,as the fig indicates,is specified by
the six torsion angles of the sugar –
phosphate backbone and the torsion
angle describing the orientation of the
base around the glycosidic bond (the
bond joining the c1’ to the base.
It would seem that these seven degrees
of freedom per nucleotide would render
the polynucleotide very flexible.
Yet these torsion angles are subject to a
variety of internal constraints that
greatly restrict their rotational freedom.
The rotation of a base around its
glycosidic bond (angle χ) is greatly
hindered.

18. THE STERICALLY ALLOWED ORIENTATIONS OF PURINE AND PYRIMIDINE BASES W.R.T.THEIR
ATTACHED RIBOSE UNITS;IN B-DNA,THE NUCLEOTIDE RESIDUES ALL HAVE ANTI CONFORMATIONS
Purines residues have two sterically permissible orientations
known as the syn-(greek:with) and anti- (greek:against)
conformations.
Only the anti conformations of pyrimidines is stable,because in
the syn conformation ,the sugar residue sterically interferes with
the pyrimidine’s C2 substituent.
In most double helical nucleic acids, all bases are in the anti
conformation.
The exception is Z-DNA,in which the alternating purine and
pyrimidine residues are anti and syn,respectively (this is the one
reason why the repeating unit of Z-DNA is a dinucleotide).

19. The flexibility of the ribose ring itself is also
limited.
The vertex angles of a regular pentagon are 108˚,a
value quite close to the tetrahedral angle
(109.5˚),so one might expect the ribofuranose ring
to be nearly flat, however the ring substituents
are eclipsed when the ring is planar.
To relieve this crowding, which occurs even
between hydrogen atoms, the ring puckers i.e. it
becomes slightly non-planar.
In the great majority of known nucleoside and
nucleotide x-ray structures, four of the ring atoms
are co-planar to within a few hundredths of an
angstrom and the remaining atom is out of this
plane by several tenths of an angstrom.
The out of plane atom is almost always c2’ and c3’.

20. The two most common ribose conformations are known as C3’-endo and C2’-
endo;”endo” (greek: end on, within) indicates that displaced atom is on the same
side of the ring as C5’.
The ribose pucker is conformationally important in nucleic acids because it
governs the relative orientations of phosphate substituents to each ribose residue.
In fact, B-DNA has the C2’-endo conformation, whereas A-DNA is C3’-endo
In Z-DNA, the purine nucleotides are all c3’-endo and the pyrimidine nucleotides
are C2’-endo.

21. DNA STRUCTURE
DNA is a molecule
duplex i.e consists of two
chains arranged in a
antiparallel manner and
with nitrogenous bases
facing each other.
In a three-dimensional
there are three different
levels
Primary
Secondary
Tertiary

22. SUMMARY OF PRIMARY,SECONDARY AND
TERTIARY STRUCTURES
Primary structure:
 Sequence of nucleotide chains. It is in these channels where the genetic
information, and because the skeleton is the same for all ,the difference in the
information lies in the different sequence of nitrogenous bases. This sequence has
a code, which determines an information or otherwise, as the order of the bases.
Secondary structure:
 It is a double helix structure. Can explain the storage of genetic information and
the mechanism of DNA replication. It was postulated by Watson and Crick, based
on X-ray diffraction that Franklin and Wilkins had made, and the equivalence of
bases Chargaff's postulated, whereby the sum of adenines more guanines is equal
to the sum of thymine more cytosine.
It is a double strand, right-handed or left-handed, depending on the DNA. Both
chains are complementary, as adenine and guanine in a chain are joined,
respectively , to thymine and cytosine on the other. Both chains are antiparallel,
then the 3 'end of one faces the 5' end of the counterpart.
 There are three models of DNA. The DNA of type B is the most abundant and is
discovered by Watson and Crick.

23. Tertiary structure:
 Refers to how DNA is stored in a confined space to form the
chromosomes. Varies depending on whether the organisms
prokaryotes and eukaryotes:
 In prokaryotes the DNA is folded like a super-helix, usually in circular
shape and associated with a small amount of protein.
 In eukaryotes, since the amount of DNA from each chromosome is very
large, the packing must be more complex and compact, this requires
the presence of proteins such as histones and other proteins of non-
histone nature (protamines).

25.  PRIMARY STRUCTURE
 A single DNA chain is a long thread
like molecule made up of a large no. of
deoxyribonucleotides.
 The backbone of primary structure
consists of deoxyribose linked by
phospho-diester bridges.
 The phospho-diester bond are formed
between 3’- and 5’-of the successive
sugar molecules.
 The 3’OH group of deoxy-pentose of
one nucleotide is joined to 5’OH group
of deoxy-pentose of the adjacent
nucleotide through a phosphate group.
 This way a long unbranched chain is
formed which has the polarity a 5’end
and a 3’end are free(phosphate groups
are free without the phospho-diester
linkage not attached to other
nucleotides).
LEVELS OF STRUCTURE OF DNA

26. SECONDARY STRUCTURE OF DNA

27.  Two antiparallel
polynucleotide chains
wound around the same
axis.
 Sugar phosphate chains
wound around the
periphery.
 Bases A,T,G and C occupy
the core , forming A:T and
G:C Watson-Crick base pairs.
 The DNA double helix is held
together mainly by-
Hydrogen Bonds.
SECONDARY STRUCTURE OF DNA

28. HYDROGEN BOND
 A chemical bond in
which a hydrogen
atom of one
molecule is
attracted to an
electronegative
atom of another
molecule (especially
a nitrogen, oxygen
or fluorine atom).

29. EVENTS LEADING TO THE DNA STRUCTURE
 In 1953,James Watson and
Francis Crick discovered
the double helical
structure of DNA.
 The scientific framework
for their breakthrough
was provided by other
scientists including
-Linus Pauling
-Rosalind Franklin
-Erwin Chargaff

30. LINUS PAULING MODEL OF DNA

31. X-RAY CRYSTALLOGRAPHY
 X-Ray diffraction study of DNA by
W.T.Astbury (1940s) indicated that DNA
is a polynucleotide chain , where
successive nucleotides occur at 3.4 A.
 Franklin (1952) observed DNA to be a
helix.
 Wilkins and Franklin (1953) obtained
very fine x-ray diffraction pictures of
DNA which were immediately made
available to Watson and Crick.
 Piecing together all the previous
information ,Watson and Crick(1953)
came to the conclusion that DNA was
made of two anti-parallel helical
chains held together by hydrogen
bonds created between their nitrogen
bases.

33. ERWIN CHARGAFF’S EXPERIMENT
 It was assumed the four bases;
A,G,C and T were in a repeating
tetranucleotide
configuration.
 Therefore , there should be the
same amount of A,G,C and T in
any molecule of DNA from any
source.
 Chargaff carefully determined
the exact percentages of
nucleotides in DNA from several
sources.
 %A=%T and %G=%C.
 However %AT did not equal to
%GC.
 This observation became known
as Chargaff Rule.

34. WATSON-CRICK MODEL OF DNA

35. BASE PAIRING
It is a pairing formed in the
DNA double helix between
purine of one strand and
pyrimidine of the second
strand.
Base pairing is specific with
adenine lying opposite
thymine and cytosine
occurring opposite guanine.
It can accommodate neither
two purines , nor two
pyrimidines.

36.  According to Watson and Crick, a DNA molecule
consists of two polynucleotide chains wrapped
helically around each other,with the sugar
phosphate chains on the outside and purines
and pyrimidines on the inside of the helix.
 The two chains are spirally coiled around a
common axis in a regular manner to form a
double helix.
 The double helix is of constant diameter of 2
nanometers(nm) or 20 angstroms and has a
major groove, about 20 angstroms wide and the
minor groove about 12 angstroms wide
alternately.
 One complete spiral helix is 34 angstroms long
and has 10 base pairs.
 The bases face the interior of the double helix
and are stacked 3.4 angstroms apart.
WATSON AND CRICKS MODEL OF DNA

37.  The sugar phosphate component forms the
backbone on the outside.
 The two strands run antiparallel.one strand
has phosphodiester linkage in the 3’ -5’
direction, while the other strand has
phosphodiester linkage in the 5’-3’ direction.
 The helix is generally right handed that is it
runs clockwise looking along the helix axis.
 The two strands are held together by
hydrogen bonds between specific base pairs
of purines and pyrimidines. The hydrogen
bond between purines and pyrimidines are
such that adenine can bond only to thymine
by two hydrogen bonds and guanine can
bond only to cytosine by three hydrogen
bonds.
 The specificity of the kind of hydrogen bonds
that can be formed assures that for every
adenine in one chain there will be thymine in
the other and for every guanine in one chain
there will be cytosine in the other. Thus the
two chains are complementary to each other.

38. DOUBLE HELIX-RIGHT HANDED AND LEFT HANDED COILING
 The double helix is a spiral right-
handed, that is, each of the nucleotide
chains turn right, this can be verified
if we look at the segment (a), where
the threads move upwards and
eventually turn right .
 If the two strands rotate clockwise it
is said that the double helix is right-
handed, and if they turn towards left,
left-handed (this form may appear in
helices alternatively because of
conformational changes in DNA).
 But the most common conformation
adopted by the DNA double helix is
right-handed, turning every couple of
bases on the previous approximately
36 º.

39.  When the two DNA strands are rolled over each other (either
left or right), cracks are formed between a thread and the
other, exposing the sides of the nitrogenous bases inside.
 In the most common conformation DNA adopts, because of
the angles between the sugars of both strands of each pair of
nitrogenous bases, appears two types of cracks around the
surface of the double helix: one, the cleft or major groove,
which is 22 Å (2.2 nm) wide, and the other, the minor groove,
which is 12 Å (1.2 nm) wide.
 The major groove is wider than the minor groove in
DNA;and many sequence specific proteins interact in the
major groove.
 The N7 and C6 groups of purines and the C4 and C5 groups of
pyrimidines,face into the major groove.
 Thus, they can make specific contacts with amino acids in
DNA binding proteins.
 Thus specific amino acids serve as H-bond donors and
acceptors to form H-bonds with specific nucleotides in
DNA.
 H-bond donors and acceptors are also in the minor groove
and indeed some proteins bind specifically ,in the minor
groove.
 Base pairs stack with some rotation between them.

40. MAJOR HELICAL CONFORMATIONS OF DNA
Most of the biologically active DNA exists in Watson-Crick form . This is the B-form of the
DNA.The double helix is able ton assume other forms depending upon varying environmental
conditions.
 6 MORPHOLOGICAL FORMS OF DNA
 A,B,C,D,E and Z.
 A-DNA: Right handed helix with 11 base pair turns . It is the dehydrated form which occurs in the
environment richer in Na+ and less of water.
 B-DNA: Watson and Crick’s model of DNA is the common form of DNA found in organisms . It is a
right handed helix with each turn of spiral having 10 base pairs . It occurs under salt
concentration and high degree of hydration.
 C-DNA: Right handed helix with 9 base pairs per turn.
 D-DNA: Right handed helix with 8 base pairs per turn.
 E-DNA: Form adapted by synthetic DNA lacking guanine . There are 7½ base pairs per turn.
 C-DNA and E-DNA are seen under special environmental conditions and have slightly different
conformation so do not occur in vivo.
 Z-DNA: Left handed helix with zigzag back of sugar phosphate residues and 12 base pair per turn
of helix . It is the skinniest DNA, with only one groove and is stabilized by high salt
concentration.

41. 3 MAJOR FORMS OF DNA

42. CONFORMATIONS OF A,B and Z DNA
DNA exists in many conformations. However,
in living organisms have only been observed
conformations A-DNA, B-DNA and Z-DNA.
The conformation DNA adopts depends on its
sequence, the amount and direction of super
coiling that show the presence of chemical
modifications on the bases and conditions of
the solution, such as the concentration of
ions of metals and polyamines.
In the three conformations, the form "B" is
the most common conditions in the cells.
The two DNA double helices alternatives
differ in their geometry and dimensions.

43. Under dehydrating conditions-DNA undergoes a reversible
conformational change to A –DNA.
A-DNA ,which forms a wider and flatter right –handed helix than does
B-DNA.
A-DNA has 11.6 bp per turn and a pitch of 34 Angstroms, which gives it
an axial hole.
A-DNA’s most striking feature ,however, is that the planes of its base
pairs are tilted 20 degree w.r.t the helix axis.
Since the axis does not pass through its base pairs, A –DNA has a deep
major groove and a very shallow minor groove ;it can be described as
a flat ribbon wound around a 6-Angstroms diameter cylindrical hole.

44.  In B DNA ,the bases occupy the core of
the helix while the sugar phosphate
backbones wind around the outside,
forming the major and minor grooves.
Only the edges of the base pairs are
exposed to the solvent.
 The “ideal” B DNA helix has 10 base pairs
(bp) per turn (a helical turn of36 degree
per bp) and, since the aromatic bases
have van der waal’s thickness of 3.4
Angstroms and are partially stacked on
each other, the helix has a pitch (rise per
turn )of 34 Angstroms.
 The helix of Z DNA has 12 BASE pairs per
turn, a pitch of 45 Angstroms ,a deep
minor groove, and no discernible major
groove.
 Z-DNA, therefore resembles a left-
handed drill bit in appearance.

49. SIMILARITIES BETWEEN Z-DNA and B-DNA.
 Both are double helical.
 In both DNAs ,two polynucleotide strands of double helix are antiparallel.
 Both forms exhibit G triple bond C PAIRING.
DIFFERENCES BETWEEN Z-DNA and B-DNA.
 Z-DNA has left-handed helical sense, while B-DNA has right handed helicalsenses
 The phosphate back bone of Z-DNA follows a zigzag course ,while in B-DNA this
backbone is regular.
 In Z-DNA , the adjacent sugar residues have opposite orientation, while in B-
DNA , they have same orientation. Due to this, the repeating unit is a
dinucleotide in Z-DNA as against a mononucleotide unit in B-DNA.
 In Z-DNA , one complete helix(i.e. a twist through 360˚ has twelve base pairs or
six repeating dinucleotide units, while in B-DNA one complete helix has only 10
base pairs or 10 repeating units.
 In Z-DNA, one complete turn of helix is 45˚ long, while in B-DNA it is 34
Angstroms long.

50. DENATURING AND ANNEALING OF DNA
The DNA double strand can be denatured if heated (95 C) or treated
with chemicals.
 AT regions denatures first (2H bonds)
 GC regions denatures last (3H bonds)
DNA denaturation is a reversible process , as DNA strands can be re-
annealed if cooled.
This process can be monitored using the hyperchromicity (melting
profile).
Hyperchromicity:
 It is used to moniter DNA denaturation and annealing.
 It is based on the fact that single stranded(SS) DNA gives higher
absorption reading than double stranded (DS) at wavelength 260
nm.

51. DENATURATION
 The strands of the DNA double helix are held
together by hydrogen bonding interactions
between the complementary base pairs. Heating
DNA in solution easily breaks these hydrogen bonds,
allowing the two strands to separate—a process
called denaturation or melting.
 The two strands may reassociate when the solution
cools, reforming the starting DNA duplex—a
process called renaturation or hybridization.
 These processes form the basis of many important
techniques for manipulating DNA. For example, a
short piece of DNA called an oligonucleotide can be
used to test whether a very long DNA sequence has
the complementary sequence of the
oligonucleotide embedded within it.
 Using hybridization, a single-stranded DNA
molecule can capture complementary sequences
from any source. Single strands from RNA can also
reassociate. DNA and RNA single strands can form
hybrid molecules that are even more stable than
double-stranded DNA.
 These molecules form the basis of a technique that
is used to purify and characterize messenger RNA
(mRNA) molecules corresponding to single genes.

52. ULTRAVIOLET ABSORBPTION
• DNA melting and reassociation can be monitored by
measuring the absorption of ultraviolet (UV) light at a
wavelength of 260 nanometers (billionths of a metre). When
DNA is in a double-stranded conformation, absorption is
fairly weak, but when DNA is single-stranded, the unstacking
of the bases leads to an enhancement of absorption called
hyperchromicity(melting profile). Therefore, the extent to
which DNA is single-stranded or double-stranded can be
determined by monitoring UV absorption.

54. MELTING POINT CURVE : Tm IS PROPORTIONAL TO %GC

56. BOUYANT DENSITY OF DNA
It is the density of the
solution at which the
DNA feels no net force
during centrifugation is
called its buoyant
density.
This is the density in
the density gradient
where that particular
DNA molecule will form
bands as it stops going
up or down.

57. • Under constant conditions (usually 25˚C in ceasium chloride at neutral
pH ) the buoyant density of DNA is related to the GC content.
• The fractionation of Phaseolus aerus DNA has a buoyant density of
1.695g/cm³ and that of E.coli DNA has a buoyant density of 1.710 g/cm³in
ceasium chloride.
• Most nuclear DNAs from higher plants have buoyant density within the
range 1.69-1.71 g/cm³.
• However the presence of 5-metyl-cytosine serves to reduce the density
slightly , thereby giving rise to an under estimate of the GC content.
• In general, 1% methylation decreases the buoyant density by 1 mg/cm³.
• Certain sequences of bases may also distort the relationship between
the base composition and buoyant density.
• Furthermore , ssDNA is denser than dsDNA of similar base composition
by approximately 0.015 g/cm³ and under alkaline conditions the density
is increased by 0.06 g/cm³

58. FACTORS AFFECTING BUOYANT DENSITY
• Buoyant Density of DNA depends on the following factors:
• Nature of the ceasium chloride,
• Presence of heavy metals or DNA binding dyes,
• The pH, and
• The temperature.