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Signaling Part 2

1) DNA replication is a highly accurate process that must copy genetic information each time a cell divides. Even one mistake per million base pairs could lead to thousands of errors. 2) Early experiments on replication models by Meselson and Stahl using nitrogen isotopes demonstrated that replication occurs in a semiconservative manner, with each parental DNA strand serving as a template to replicate a new partner strand. 3) Replication occurs via bi-directional replication forks moving along DNA. On the leading strand, replication is continuous while the lagging strand involves discontinuous replication in short Okazaki fragments that are later joined.

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Wissam Alwazani
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41 views27 pages

Signaling Part 2

1) DNA replication is a highly accurate process that must copy genetic information each time a cell divides. Even one mistake per million base pairs could lead to thousands of errors. 2) Early experiments on replication models by Meselson and Stahl using nitrogen isotopes demonstrated that replication occurs in a semiconservative manner, with each parental DNA strand serving as a template to replicate a new partner strand. 3) Replication occurs via bi-directional replication forks moving along DNA. On the leading strand, replication is continuous while the lagging strand involves discontinuous replication in short Okazaki fragments that are later joined.

Uploaded by

Wissam Alwazani
Copyright
© © All Rights Reserved
We take content rights seriously. If you suspect this is your content, claim it here.
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Download as PDF, TXT or read online on Scribd
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DNA Replication

1
DNA
Replication
DNA
Reverse Transcription
Transcription
Translation
RNA RNA protein
Replication

2
Genetic Information Must Be Accurately Copied Every
Time a Cell Divides

• Replication has to be extremely accurate:

• 1 error/million bp leads to 6400 mistakes


every time a cell divides, which would be
catastrophic.

• Replication also takes place at high speed:

• E. coli replicates its DNA at a rate of 1000


nucleotides/second.
Replication Models:

• Conservative replication model

• Dispersive replication model

• Semiconservative replication
Meselson and Stahl’s Experiment

• Two isotopes of nitrogen:

• 14N common form; 15N rare heavy form

• E. coli were grown in a 15N media first, then


transferred to 14N media.

• Cultured E. coli were subjected to equilibrium


density gradient centrifugation.
Meselson and Stahl (1958)

7
Modes of Replication

• Replicons: units of replication

• Replication origin

• Theta replication: circular DNA, E. coli; single


origin of replication forming a replication fork,
usually a bidirectional replication

• Rolling-circle replication: virus, F factor of


E. coli; single origin of replication
Linear Eukaryotic Replication

• Eukaryotic cells; thousands of origins; a typical


replicon: 200,000 ~ 300,000 bp in length
Uni or bidirectional
Replication forks move in one or opposite directions
◼
Linear Eukaryotic Replication

• Requirements of replication:

• A template strand

• Raw material: nucleotides

• Enzymes and other proteins


Linear Eukaryotic Replication

• Direction of replication:

• DNA polymerase add nucleotides only to the


3′ end of a growing strand.

• The replication can only go 5′→3′.


Linear Eukaryotic Replication
• Direction of replication:

• Leading strand: undergoes continuous


replication

• Lagging strand: undergoes discontinuous


replication

• Okazaki fragment: the discontinuously


synthesized short DNA fragments
forming the lagging strand
Semi-discontinuous replication
Anti parallel strands replicated simultaneously
❑ Leading strand synthesis continuously in 5’– 3’
❑ Lagging strand synthesis in fragments in 5’-3’
Core proteins at the replication fork

Topoisomerases - Prevents torsion by DNA breaks


Helicases - separates 2 strands
Primase - RNA primer synthesis
Single strand - prevent reannealing
binding proteins of single strands
DNA polymerase - synthesis of new strand
Tethering protein - stabilises polymerase
DNA ligase - seals nick via phosphodiester
linkage
The mechanism of DNA replication
Arthur Kornberg, a Nobel prize winner and
other biochemists deduced steps of
replication
– Initiation
• Proteins bind to DNA and open up double helix
• Prepare DNA for complementary base pairing
– Elongation
• Proteins connect the correct sequences of
nucleotides into a continuous new strand of DNA
– Termination
• Proteins release the replication complex
Supercoiled DNA relaxed by gyrase & unwound by helicase + proteins:

5’ SSB Proteins
Okazaki Fragments
ATP
1
Polymerase III 2 Helicase

Lagging strand 3 +
Initiator Proteins

3’
primase base pairs

Polymerase III 5’
RNA primer replaced by polymerase I

& gap is sealed by ligase

Leading strand
RNA Primer

3’ 23
DNA ligase seals the gaps between Okazaki fragments with a
phosphodiester bond

24
Model of DNA replication

25
Model of DNA replication

26
See you in the next lecture
With my best wishes

THANK YOU
27

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