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6.2: Dna Replication

DNA replication follows the semi-conservative model where each new DNA molecule contains one original parental strand and one newly synthesized strand. It involves the unwinding of the DNA double helix by the enzyme helicase. The separated strands then act as templates for DNA polymerase to synthesize new complementary strands in the 5’ to 3’ direction. This results in the formation of a leading continuous strand and a lagging discontinuous strand made of Okazaki fragments that are later joined together by DNA ligase. Key enzymes involved are DNA polymerase, primase, ligase, helicase and topoisomerases. Meselson-Stahl experiment provided evidence supporting the semi-conservative model of DNA replication.

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56 views43 pages

6.2: Dna Replication

DNA replication follows the semi-conservative model where each new DNA molecule contains one original parental strand and one newly synthesized strand. It involves the unwinding of the DNA double helix by the enzyme helicase. The separated strands then act as templates for DNA polymerase to synthesize new complementary strands in the 5’ to 3’ direction. This results in the formation of a leading continuous strand and a lagging discontinuous strand made of Okazaki fragments that are later joined together by DNA ligase. Key enzymes involved are DNA polymerase, primase, ligase, helicase and topoisomerases. Meselson-Stahl experiment provided evidence supporting the semi-conservative model of DNA replication.

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a200812
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CHAPTER 6 : BIOLOGY 1 SB015

EXPRESSION OF BIOLOGICAL INFORMATION SEMESTER 1


SESSION 2022/2023

6.2 : DNA REPLICATION

LEARNING OUTCOMES
a) Explain semi-conservative replication of DNA.
b) Explain the enzymes and proteins involved in
DNA replication.
c) Explain the mechanism of DNA replication and
the enzymes involved.
6.2 DNA REPLICATION
Watson and Crick’s Hypothesis

Both polynucleotide
First step in replication
strands of DNA is
is separation of 2 DNA
complementary to
strands
each other
6.2 DNA REPLICATION

Parental / old strands

Each parental strand act


as template to produce
new complementary
strands; according to
base-pairing rule

New strands
6.2 DNA REPLICATION

Two identical DNA strands


are formed from one DNA
strand; each daughter DNA
molecule consists of one
parental strand and one
new strand.
REVIEW OF DNA REPLICATION

2 daughter DNA
New strands
Parental strands
(act as template)
Three Alternative Models of DNA Replication

1. Semi-conservative model

- Two parental strands of


parental molecule separate,
each functions as a
template for synthesis a
new, complementary strand.
- Each DNA consist of
one old strand and
one new strand.
Three Alternative Models of DNA Replication

2. Conservative model

- Two parental strands


reassociate after acting as
templates for new strands,
thus restoring the parental
double helix.
- Each DNA molecule
consist of two old strands
and two new strands.
Three Alternative Models of DNA Replication

3. Dispersive model

- Each strand of both


daughter molecules
contain a mixture of
old and newly
synthesized DNA.
MESELSON AND STAHL’S EXPERIMENT

• Matthew Meselson & Franklin Stahl (1950)


devised experiments that tested the three
hypotheses.
• Their experiments supported semi-
conservative model.
MESELSON AND STAHL’S EXPERIMENT

Materials: 1) Heavy isotope of nitrogen 15N


2) Lighter isotope of nitrogen 14N

Bacteria: Escherichia coli


MESELSON AND STAHL’S EXPERIMENT
MESELSON AND STAHL’S EXPERIMENT

CONCLUSION

- DNA replication follows


semi-conservative model.

- The result eliminated


the conservative and
dispersive models.
REVIEW OF DNA REPLICATION

● DNA replication is
semi-conservative.
● Each strand acts

as template.
● Free DNA nucleotide

join up to exposed
bases by specific
base pairing.
Basic Rules of DNA Replication

1. Semi-conservative
2. Starts at the origin
3. Synthesis always in the 5’-3’ direction
(new strand)
4. Can be unidirectional or bidirectional
5. Primers required
DNA Replication

⮚ 5 enzymes and 1 protein are involved:

1. Helicase
2. Single-strand binding protein (SSB protein)
3. Topoisomerase
4. Primase
5. DNA polymerases
6. DNA ligase
He said “ Please don’t do lies”

Single strand DNA primase


Helicase binding protein
He said Please

DNA DNA DNA ligase


polymerase III polymerase I
D on’t Do Lies
Helicase

Helicase are the enzymes that unwind the


double helix DNA at the replication forks, and
separate two parental strands to become single
strand.
Single-strand binding proteins

Single-strand binding proteins are the


proteins that hold the separated parental
strands apart while they act as template.
Topoisomerase

Topoisomerase are the enzymes that helps


relieve the strain caused by unwinding.
Primase

Primase are the enzymes that catalyze the


addition of RNA nucleotides that
complementary to the template to form RNA
primer.
DNA polymerase

DNA polymerase are the enzymes that


catalyze the addition of DNA nucleotides.

There are two types of DNA polymerase:

• DNA polymerase III


• DNA polymerase I
DNA polymerase III

DNA polymerase III adds free DNA nucleotides


to the exposed bases by specific base pairing
at 3’ end of RNA primer.

RNA
primer

3’
5’

DNA polymerase III


DNA polymerase I

DNA polymerase I removes the RNA primer


and replace with DNA nucleotides
DNA ligase

DNA ligase are the enzymes that catalyzed the


joining of sugar-phosphate backbone of all
Okazaki fragments together to form a
continuous strand

DNA ligase
DNA Replication
DNA REPLICATION PROCESS

Step 1: Unwinding of strand


1. DNA replication begins at origin of replication
2. Enzyme helicase unwind the double helix strand of
DNA and separate two parental strand to become
single strand.
3. Forming replication bubble. Replication fork is a Y
4. At each end of replication bubble is shape region where
parental strands of DNA
a replication fork. are being unwound
DNA REPLICATION PROCESS

DNA templates

Step 2: Strand acts as template


5. When parental strands of DNA are being unwound,
both DNA strands act as template
6. Single-strand binding proteins hold the separated
parental strands apart while they act as template.
7. SSB protein bind to the unwound DNA strands and
stabilize the single-stranded DNA.
DNA REPLICATION PROCESS

DNA templates

Step 2: Strand acts as template


8. The unwinding of double helix causes tighter
twisting and strain ahead of replication fork
9. Topoisomerase helps relieve the strain caused by
unwinding.
DNA REPLICATION PROCESS
Step 3: Synthesis of RNA primer
10. Primase catalyze the addition of RNA nucleotides that
complementary to the template to form RNA primer.

RNA primer is a short


segment of RNA
which consist of 5 to
10 nucleotides
Why Priming ?
• This is because DNA polymerase cannot initiate the
synthesis of new DNA strands.
• They can ONLY add DNA nucleotides to the end of
an existing chain that is base-paired with the
template strand.
• The existing chain refer to RNA primer.

RNA
primer
DNA REPLICATION PROCESS

Step 4: DNA nucleotides join up to the exposed bases


by specific base pairing
11. DNA polymerase III adds free DNA nucleotides to the
exposed bases by specific base pairing at 3’ end of
RNA primer.

RNA primer

3’
5’

DNA polymerase III


DNA REPLICATION PROCESS

Step 4: DNA nucleotides join up to the exposed bases


by specific base pairing
12. DNA polymerase III catalyze the elongation of new DNA
strand at replication fork ONLY in 5’ 3’ direction.

3’ 5’
DNA REPLICATION PROCESS
Step 5: Formation of leading and lagging strands from
5’ to 3’
13. Leading strand synthesized continuously along the
template strand which is elongates towards the
replication fork.
14. Lagging strand synthesized discontinuously along the
template strand which is elongates in the direction away
from the replication fork.
DNA REPLICATION PROCESS
Step 5: Formation of leading and lagging strands from
5’ to 3’
15. Lagging strand consist of short fragments called
Okazaki fragments.
SYNTHESIS OF LEADING STRAND
SYNTHESIS OF LAGGING STRAND
Reiji Okazaki
SYNTHESIS OF LEADING & LAGGING STRAND

3'

5'
DNA REPLICATION PROCESS

Step 5: Formation of leading and lagging strands from


5’ to 3’
16. DNA polymerase I removes the RNA primer and
replace with DNA nucleotides
DNA REPLICATION PROCESS

Step 5: Formation of leading and lagging strands from


5’ to 3’
17. DNA ligase catalyzed the joining of sugar-phosphate
backbone of all Okazaki fragments together to form a
continuous strand

DNA ligase
Lagging strand
DNA REPLICATION PROCESS
18. Two identical copies of the original DNA are
produced.
SUMMARY

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