DNA Structure and

Replication

Mamatha Garige, Ph.D.

Nucleic
Acidsacids are long, linear polymers constructed from
Nucleic
four types of monomers.
Each monomer consists of a sugar, a phosphate, and a base.
The sequence of the bases is the information content of the
nucleic acid.

DNA and RNA

The sugar component of deoxyribonucleic acid (DNA) is
deoxyribose, a ribose in which the 2-hydroxyl is replaced with
a hydrogen.
Ribonucleic acid (RNA) contains the sugar ribose.
Two of the bases are purines (adenine and guanine) and two are
pyrimidines (cytosine and thymine (DNA) or uracil (RNA)).

Base
s

Nucleoside A base bound to a sugar is called a nucleoside.

The nucleosides of DNA are
deoxyadenosine, deoxyguanosine,
deoxycytidine, and deoxythymidine.
The nucleosides of RNA are adenosine,
guanosine, cytidine, and uridine.
In all cases, the C-1 of the sugar is
attached to the N-9 of the purine or the
N-1 of the pyrimidine.

Nucleotid
e

A nucleotide is a nucleoside with one or more

phosphoryl groups attached.
Nucleoside triphosphates are the
building blocks of DNA and RNA.

The backbones of DNA and

RNA consist of the sugars
linked by phosphodiester
bonds between the 3hydroxyl of one ribose sugar
and the phosphoryl group
linked to 5-hydroxyl of an
adjacent ribose sugar
molecule.
Bases are attached to
carbon atom 1 in the sugar.

General features of DNA are:

1. DNA molecules consist of two chains
of opposite directionalityone strand
runs in the 5 to 3 direction and the
other in the 3 to 5 direction
intertwined to form a right-hand
double helix.
2. The sugar-phosphate backbones are
on the outside of the helix, whereas
the bases are inside the helix.
3. The bases are perpendicular to the
axis of the helix with adjacent bases
separated by 3.4 .
4. The helix is approximately 20 wide.
5. Adenine always hydrogen bonds with
thymine, cytosine hydrogen bonds
with guanine.

Watson and Crick Base Pairing

DNA replication is a semiconservative process (Meselson

and Stahl)
Because of the base pairing rules,
the sequence of one strand
determines the sequence of the
partner strand.
The two strands can be separated
and complementary sequences
synthesized to generate two
identical daughter strands.
Because the two daughter helices
have one parent strand and one
newly synthesized strand, the
replication process is called
semiconservative replication.

In the cell, the most commonly

seen form of DNA double helix is
called the B form.
The double helix can also exist in
an A form, which is shorter and
wider than the B form with the
bases at an angle rather than
perpendicular to the helix axis.
The A form is seen in RNA double
helices and in RNA-DNA hybrid
helices, structures observed in
transcription and RNA processing.

The double helix can also form Z

DNA. Z DNA is left-handed and the
backbone is zigzagged, accounting
for the name Z DNA.

DNA Packaging
There are 3.6 meters of DNA in a human cell, packaged
into 46 chromosomes.
Supercoiling accounts for some of the compaction of the
DNA, but further compaction occurs by binding certain
proteins called histones to the DNA
Chromatin is the entire complement of a cells DNA
and its associated proteins.
Histones are highly basic proteins that are components of
chromatin.
Octet comprised of two copies each of histones H2A, H2B,
H3, H4 and 200 bp of DNA comprise a nucleosome.
Nucleosomes are joined by linker DNA, to which histone H1
binds, so that the histone-DNA complex has the appearance
of beads on a string.
Digestion of the linker DNA yields the nucleosome core
particle.

The Compaction of DNA into a Eukaryotic Chromosome

The mammalian cell

cycle
Rapid growth and
preparation for
DNA synthesis

phas
e

G1
phase

DNA replication
and
histone
synthesis

G2
M

phase

phase

Growth and
preparation
for
cell division

Mitosis: replicated chromosomes

are segregated into daughter

DNA
Replication

The reaction catalyzed by DNA polymerase is:

Key characteristics of DNA synthesis are:

1. Four deoxynucleoside triphosphates and Mg2+ are required.
2. A template strand is used to direct DNA synthesis.
3. A primer from which the new strand grows must be present.
4. Many DNA polymerases have nuclease activity that allows
for the removal of mismatched bases.

Other enzymes involved in DNA

Replication
Helicases, powered by ATP hydrolysis, separate the
strands of the double helix to make the DNA available for
DNA polymerase.
Defects in the helicase can result in Werner syndrome, a
pathological condition characterized by premature aging
Topoisomerases are enzymes that induce or eliminate
supercoils.
Type 1 topoisomerases relax supercoils while type 2
topoisomerase, such as DNA gyrase in E. coli, introduce
supercoiling.

A special type of RNA polymerase called primase

synthesizes short RNA (10 nucleotides) complementary
to a DNA strand that then serves as a primer for DNA
synthesis.

 The site of replication is called the replication fork.

The fork moves in one direction, demonstrating that both
strands are copied simultaneously.
However, all DNA polymerases synthesize DNA only in the 5
3 direction.
At the replication fork, one strand is synthesized
continuously in the 53 direction.
The other strand is synthesized discontinuously as small
pieces called Okazaki fragments in the 53 direction.

DNA polymerase 35
exonuclease activity
removes the RNA primer
and replaces the sequence
with DNA.
DNA ligase joins the
fragments to yield an
intact strand.

The free ends of linear DNA molecules present two

biochemical difficulties.
1.They are susceptible to damage by nucleases.
2.Due to the nature of DNA synthesis, one strand will
shorten upon each round of DNA synthesis.
The ends of the chromosomes are called telomeres. The
longer of the two strands is rich in guanine. The stretch of
guanine-rich single-strand DNA can form a loop structure
to protect the end of the chromosome.

The G-rich longer strand of the telomere can be

maintained by the enzyme telomerase.
Telomerase contains an RNA template that it uses to
extend the leading strand.
In rapidly dividing cells, including cancer cells,
telomeres must be maintained by the telomerase to
prevent shortening of the lagging, which would lead to
cell death.
High telomerase activity is a characteristic of cancer
cells.

Multiple origins or
replication

Single origin of
replication

Bases can be damaged by Oxidizing agents,

alkylating agents
and light (mutagens)

Deamination

Repair Mechanisms
Photoreactivation repair:
photoreactivating enzyme absorbs energy
from UV radiation (region closer to blue
light) and cleaves the pyrimidine dimers.
Base excision repair.
Nucleotide excision repair.
Recombination repair.

Base-excision
Repair

Nucleotide excision
repair

Recombinatio
n Repair