DNA replication by moazzam

Replication
of DNA
By MOAZZAM ALI SHAHID
1/20/2015 1
DNA Replication By Moazzam Ali
Shahid

Nucleic acids are required for
the storage and expression
of genetic information.
There are two types of
nucleic acids
• DNA
• RNA
Nucleic Acid
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 The DNA double helix has
two strands twisted
together.
 (In the rest of this unit we
will look at the structure
of one strand.)
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N.Base
phosphate
deoxyribose

 The backbone of
the molecule is
alternating
a sugar, parts The
teeth are
nitrogenous.
phosphate
deoxyribose
bases
phosphate
deoxyribose
bases

 Remember, DNA has
two strands that fit
together something
like a zipper.
 The teeth are the
nitrogenous bases but
why do they stick
together?

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 When making
hydrogen bonds,
cytosine always pairs
up with guanine,
 And adenine always
pairs up with
thymine.
 (Adenine and
thymine are shown
here.)
C
C
C
C
N
N
O
O
C

One deoxyribose together
with its phosphate and base
make a nucleotide.
C C
C
O
Phosphate
O
O -P O
O
O
O -P O
O
O
O -P O
O
O
Nitrogenous
base
Deoxyribose
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 Each cell has about 2
m of DNA.
 The average human
has 75 trillion cells.
 The average human
has enough DNA to go
from the earth to the
sun more than 400
times.
 DNA has a diameter of
only 0.000000002 m.
The earth is 150 billion m
or 93 million miles from
the sun.

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DNA Replication(
sarib_virgo@yahoo.com) 19

Three Models of DNA Replication
Conservative model
Parent molecule remains the same
Completely new copy of the double helix is made
Semiconservative model
Parent strands separate and serve as templates for new
strand synthesis
Hybrid molecules are made
Dispersive model
New strands contain a mixture of old molecules and
newly synthesized molecules
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 Semiconservative
Model:
1. Watson and Crick
showed: the two strands
of the parental molecule
separate, and each
functions as a template
for synthesis of a new
complementary strand.
.
Parental DNA
DNA Template
New DNA
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DNA Replication(
sarib_virgo@yahoo.com) 24

1. DNA replication occurs on template in 5’ - 3’
direction.
2. Requires RNA primer for de novo synthesis: made by
primase.
3. Asymmetric: Continuous on leading strand
Discontinuous on lagging strand
4. Okazaki Fragments on lagging strand.
5. Replication is semi-conservative.
6. Specific origins of replication. Sequence dependent.
Usually bidirectional.
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STEPS INVOLVED IN DNA REPLICATION
1. Identification of the origins of replication (ori)
2. Unwinding (denaturation) of dsDNA to provide a
ssDNA template
3. Formation of replication fork
4. Initiation of DNA synthesis and elongation
5. Formation of replication bubbles with ligation of the
newly synthesized DNA segment
6. Reconstitution of chromatin structure
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Enzymes in DNA Replication
•Helicases
•SSB
•DNA Primase
•DNA Polymerases
•DNA Ligase
•Topoisomerases
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Polymerase Functions Proof Reading
Pol alpha Contain
Primase
-
Initiate DNA -
Pol Beta Repair +
Pol gama Replicates
mDNA
+
Pol sigma Elongate
leading and
lagging strand
+
Pol eta Repair +
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DNA Polymerase in Eukaryotes

origins of DNA replication (every ~150 kb)
replication bubble
daughter chromosomes
fusion of bubbles
bidirectional replication
Origins of DNA replication on mammalian chromosomes
5’
3’
3’
5’
5’
3’
3’
5’
3’
5’
5’
3’
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3’
5’ 3’
RNA primer
newly synthesized DNA
5’
5’
DNA polymerase
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Discontinuous synthesis of DNA
3’
5’
5’ 3’
3’ 5’
Because DNA is always synthesized in a 5’ to 3’ direction,
synthesis of one of the strands...
5’
3’
...has to be discontinuous.
This is the lagging strand.
5’
3’
3’
5’
5’
3’
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3’
5’
5’ 3’
3’ 5’
5’
3’
3’
5’
5’
3’
leading strand (synthesized continuously)
lagging strand (synthesized discontinuously)
Each replication fork has a leading and a lagging strand
• The leading and lagging strand arrows show the direction
of DNA chain elongation in a 5’ to 3’ direction
• The small DNA pieces on the lagging strand are called
Okazaki fragments (100-1000 bases in length)
replication fork replication fork
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RNA primer
5’
3’
3’
5’
3’
5’
direction of leading strand synthesis
direction of lagging strand synthesis
replication fork
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5’
3’
3’
5’
3’
5’
Strand separation at the replication fork causes positive
supercoiling of the downstream double helix
• DNA gyrase is a topoisomerase II, which
breaks and reseals the DNA to introduce negative
supercoils ahead of the fork
• Fluoroquinolone antibiotics target DNA gyrases in many
gram-negative bacteria: ciprofloxacin and levofloxacin (Levaqu
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5’
3’ 5’
3’
Movement of the replication fork
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Movement of the replication fork
RNA primer
Okazaki fragment
RNA primer
5’
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newly synthesized DNA
(Okazaki fragment)5’
3’
5’
3’
DNA ligase seals the gap by catalyzing the formation
of a 3’, 5’-phosphodiester bond in an ATP-dependent reaction
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1. Why is replication necessary?
2. When does replication occur?
3. Describe how replication works.
4. Use the complementary rule to
create the complementary strand:
A---?
G---?
C---?
T---?
A---?
G---?
A---?
G---?
C---?
A---?
G---?
T---?
Replication Quiz
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1. Why is replication necessary?
So both new cells will have the correct DNA
2. When does replication occur?
During interphase (S phase).
3. Describe how replication works.
Enzymes unzip DNA and complementary
nucleotides join each original strand.
4. Use the complementary rule to
create the complementary strand:
A---T
G---C
C---G
T---A
A---T
G---C
A---T
G---C
C---G
A---T
G---C
T---A
Replication Quiz
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Components of the replication apparatus
dnaA binds to origin DNA sequence
Primasome
dnaB helicase (unwinds DNA at origin)
dnaC binds dnaB
dnaG primase (synthesizes RNA primer)
DNA gyrase introduces negative supercoils ahead
of the replication fork
Rep protein helicase (unwinds DNA at fork)
SSB binds to single-stranded DNA
DNA pol III primary replicating polymerase
DNA pol I removes primer and fills gap
DNA ligase seals gap by forming 3’, 5’-phosphodiester
bond
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