dna replication by moazzam
TRANSCRIPT
Nucleic acids are required for
the storage and expression
of genetic information.
There are two types of
nucleic acids
• DNA
• RNA
Nucleic Acid
1/20/2015 2DNA Replication By Moazzam Ali
Shahid
The DNA double helix has two strands twisted together.
(In the rest of this unit we will look at the structure of one strand.)
1/20/2015 5DNA Replication By Moazzam Ali Shahid
The backbone of the molecule is alternating
a sugar, parts The teeth are nitrogenous.
phosphate
deoxyribose
bases
1/20/2015 7DNA Replication By Moazzam Ali Shahid
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?
1/20/2015 8DNA Replication By Moazzam Ali Shahid
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
1/20/2015 14DNA Replication By Moazzam Ali Shahid
One deoxyribose together with its phosphate and base
make a nucleotide.
C C
C
OPhosphate
O
O -P O
O
O
O -P O
O
O
O -P O
O
O
Nitrogenous
base
Deoxyribose
1/20/2015 15
DNA Replication By Moazzam Ali
Shahid
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.
1/20/2015 16DNA Replication By Moazzam Ali Shahid
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
1/20/2015 20
DNA Replication By Moazzam Ali
Shahid
Semiconservative Model:
1. Watson and Crick showed: the two strands of the parental moleculeseparate, and each functions as a templatefor synthesis of a new complementary strand.
.Parental DNA
DNA Template
New DNA
1/20/2015 21
DNA Replication By Moazzam Ali
Shahid
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.
1/20/2015 25
DNA Replication By Moazzam Ali
Shahid
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
1/20/2015 26
DNA Replication By Moazzam Ali
Shahid
Enzymes in DNA Replication
•Helicases
•SSB
•DNA Primase
•DNA Polymerases
•DNA Ligase
•Topoisomerases
1/20/2015 27
DNA Replication By Moazzam Ali
Shahid
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 +
1/20/2015
DNA Replication By Moazzam Ali
Shahid 28
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’
3’1/20/2015 29
DNA Replication By Moazzam Ali
Shahid
3’
5’ 3’
RNA primer
newly synthesized DNA
5’
5’
DNA polymerase
1/20/2015 30
DNA Replication By Moazzam Ali
Shahid
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’
1/20/2015 31
DNA Replication By Moazzam Ali
Shahid
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
1/20/2015 32
DNA Replication By Moazzam Ali
Shahid
RNA primer
5’
3’
3’
5’
3’
5’
direction of leading strand synthesis
direction of lagging strand synthesis
replication fork
1/20/2015 33
DNA Replication By Moazzam Ali
Shahid
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 (Levaquin)
1/20/2015 34
DNA Replication By Moazzam Ali
Shahid
Movement of the replication fork
RNA primer
Okazaki fragment
RNA primer
5’
1/20/2015 36
DNA Replication By Moazzam Ali
Shahid
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
1/20/2015 37
DNA Replication By Moazzam Ali
Shahid
1. Why is replication necessary?
2. When does replication occur?
3. Describe how replication works.
4. Use the complementary rule tocreate the complementary strand:
A---?
G---?
C---?
T---?
A---?
G---?
A---?
G---?
C---?
A---?
G---?
T---?
Replication Quiz
1/20/2015 38
DNA Replication By Moazzam Ali
Shahid
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
1/20/2015 39
DNA Replication By Moazzam Ali
Shahid
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
1/20/2015 40
DNA Replication By Moazzam Ali
Shahid