DNA REPLICATION

LOPAMUDRA MOHAPATRA B.Sc(BIOTECH) 3rd yr ROLL NO:11303

SUBMITTED TO: TANMAYEE SAMANTARAY

CONTENTS:INTRODUCTION OF DNA: chemical composition structure of DNABASIC RULE FOR REPLICATION OF ALL

NUCLEIC ACIDSDNA REPLICATION: Initiation Elongation Termination

INTRODUCTION OF DNA:DNA: deoxy ribose nucleic acidDNA is a genetic material.DNA is a double stranded helical structure.DNA is also called as a heriditary material.

CHEMICAL COMPOSITION:

DNA isolated from variety of plants ,animals ,bacteria & virus has shown a complex form of polymeric compounds containing four monomers known as deoxy ribonucleotide monomers or deoxyribotids.

Each deoxy ribonucleotide consists of a pentose sugar ,a phosphate group & a nitrogenous base.

NITROGENOUS BASE: There are 2 nitrogen bases.i. PurineIt is of two types.1.Adenine(6-amino purine)2.Guanine(2-amino-6-oxo purine)ii. PyrimidineIt is of 3 types.3.Thymine(2,4-dioxo-5-methyl pyrimidine)4.Cytosine(4-amino-2-oxo pyrimidine)5.Uracil(2,4-dioxo pyrimidine)

STRUCTURE OF DNA:In 1953 James Watson & Hary Crick worked on the

DNA.They gave the double helical structure of DNA by

the 3 techniques.1.X-ray diffraction technique.2.Base pairing.3.AnalyticalDNA consist of two helical structure of

polynucleotide & it is a right handed double helix.The two strands of double helix are antiparallel.

Each polynucleotide chain has a sugar phosphate backbone with nitrogenous bases directed inside the helix.

The nitrogenous bases of two antiparallel polynucleotide strands are linked through hydrogen bonds.

There are 2 hydrogen bonds between A &T.There are 3 hydrogen bonds between G &C. The hydrogen bonds are not only the attractive forces

between the two polynucleotide of double helix but also serves to hold the structure together.

The two polynucleotides in a double helix are complementary.

Ten base pairs occur per turn of helix .The spacing between adjacent base pairs 10angstrom.The helix is 20angstrom in diameter.

BASIC RULE FOR REPLICATION OF ALL NUCLEIC ACID:

The prime role of any mode of replication is to produce the base sequence of the parent molecule.The specificity of base pairing-adenine with thymine & guanine with cytosine-provides the mechanism used by all replication systems.

DNA REPLICATION:DNA replication is a means to produce new molecules that have the same base sequence.Occurs during interphase of the cell cycle.DNA replication is semi-conservative • The parent DNA strand separates

into two • Each strand serves as a template for

new complementary strands• The new double helix is half original

SEMICONSERVATIVE REPLICATION OF DOUBLE SRANDED DNA:

The purpose of DNA replication is to create daughter DNA molecules that are identical to the parental molecule.

Originally,two modes of replication were suggested which differ in whether or not the two single strands of the parent molecule become rearranged after one round of replication .These were called the semiconservative & conservative modes.

http://upload.wikimedia.org/wikipedia/commons/a/a2/DNAreplicationModes.png

It is hydrogen bonded to its parental template. In the semiconservative mode, first proposed by

Watson & Crick, each parental DNA strands serve as a template for one new or daughter strand & each new strand is formed.

This mechanism ensource that genetic information will be copied faithfully at each cell division.

This process includes 3 steps.o Initiationo Elongationo Termination

The origin of replication is called as Ori C. This has two conserved regions 13 nt & 9 nt.13 nt region has 3 copies & 9 nt region has 5 copies. These regions are also called as Okazaki Fragment.

INITIATION:DnaA is a protein which will bind to the Ori C at the 9 nt regions. This binding occurs in every copy.To this DnaA complex , a complex of protein named DnaB ,will bind. That DnaA complex along with DnaB protein forms prepriming complex.The DNA has a coiled structure. This is uncoiled by a helicase named DnaB .While uncoling the knot moves forward& gradually the DNA opens.But when the coil won’t open further, at this stage a tersional stress or turgidity acts ,which is reduced to topoisomerase .Thus ,uncoiling contains & gradually , replication initiates.

ELONGATION:In this whole process DNA polymerase III is the main replicating enzymes .This enzymes has 5’->3’ & 3’->5’ exonuclease activity.RNA polymerase enzymes synthesis a RNA primer .This primer is attached to the lagging strand in the 5’->3’ site .This primer has 4-15 nt.After the primer gets attached, the DNA polymerase enzymes gets attached on the free 3’ end.The Okazaki Fragments formation takes place.The complementary strand formation takes place in the leading strand. The strand extends upto 1000 bp.

3’ 5’

RNA primase

RNA primer

After RNA primer is in place DNA nucleotides are added by DNA polymerase III.

Eventually, the segment of DNA will run into another RNA primer.

The DNA segments are called Okazaki fragments.

3’

5’

RNA primerDNA polymerase III

Okazaki fragment

Once Okazaki fragments are formed DNA polymerase I replaces the RNA primer with DNA nucleotides.

DNA ligase links the fragments together.

3’

5’

3’

5’

Parent strand

New strand

The enzymes(polymerase III holoenzymes) i.e the main replicating enzyme has 3 main subunits α,ε,θ.

The accessory subunits are γ,δ, δ’,ψ,x,β. present on template DNA α-primers are present on the core α.

ε-3’->5’ exonuclease activity.θ-function is unclear.β-helps in clamping action.γ-frameshift action.

γ-subunits helps to attach & dettach the enzyme from the lagging strands.

SSB(single stranded binding) protein gives stability & protects SSDNA.This SSB gets removed automatically.

In the lagging strand DNA replication is discontinuous

DNA is replicated in segments that become joined together.

DNA Replication (the lagging strand)

DNA replication (the leading strand)Replication is continuousThere are no fragmentsHelicase unwinds and unzips the double helixDNA polymerase III adds nucleotides in the

direction of 5’→3’DNA polymerase only works in the direction of

5’→3’

TERMINATION:For termination , first a termination point is determined . This is done by TUS protein.TUS PROTEIN:

• If replication in one side is faster than the other , then TUS is used.

• There are many B-strands in the TUS protein , which doesn’t allow the further replication of the strand.

• They are unlinked by the topoisomerase-IV.

REFERENCE: MOLECULAR BIOLOGY

by FREIFELDER

THANK YOU LOPAMUDRA MOHAPATRA