enzymes invovle in dna replication

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By: mscbiotechiti (9 month(s) ago)

vry helpful. easy to remember content. thankxx

 

By: mscbiotechiti (9 month(s) ago)

vry helpful. easy to remember content. thankxx

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Cellular & molecular biology:

Cellular & molecular biology By Mohammed razeeth.S

discussion:

discussion Enzymes involve in DNA réplication

Enzymes participation :

Enzymes participation RNA primase DNA polymerase delta DNA polymerase epsilon Exonuclease(DNA polymerase 1) DNA ligase Nuclease A Topoisomerase A gyrase Helicase

OVERVIEW:

OVERVIEW Topoisomerase Topoisomerase is required to relax th tension genered by unwinding douplex DNA. Gyrase Gyrase may be required to help unwind double helix for initiation of replication

Slide 5:

DNA polymerase DELTA Binds to 5’-3’ strands in order to be nucleotide and create leading strand DNA polymerase EPSILON Bind to 3’-5’ strands in order to create discontinuous strand called lagging strand or okazaki pices

Slide 6:

Exonuclease (DNA polymerase I) Fine and remove the RNA primer DNA ligase Adds phosphates in the remaining gap of the phosphate sugar backbone Nucleases Remove long nucleotide from the daughter strand

Slide 7:

RNA Primase Attach RNA primers to the replicating strand SSB SSB also called helix destabilizing protein. Function to maintain DNA in single strand state

Topoisomerase:

Topoisomerase Topoisomerase I cut only one strand of DNA Topoisomerase is required to relax tension general by unwinding duplex DNA The cuts usually produce 5’OH group and 3’phosphate which is covalently linked to tyrosin in the enzyme

Topoisomerase II:

Topoisomerase II Topoisomerase II is also called Gyrase. It cut the both strand. DNA Gyrase can reduce the linkage number of strands

DNA Helicase :

DNA Helicase DNA helicase have the property of being able to unwind DNA It obtain necessary energy from hydrolyze of ATP

Nuclease enzyme:

Nuclease enzyme Enzyme act to hydrolysis or breakdown of polynucleotide chain Nuclease will act either 3’-5’ end f this nuclease Nuclease have two types Exonuclease endonuclease

Slide 15:

Exonuclease Nuclease which begin attacks from a free end of polynucleotide enzyme Endonuclease Nuclease enzyme also attacks one of the two side of phosphodiester linkage but the react with only those bonds that occur with in the interior of polynucleotide chain.

Discovery of DNA polymerase:

Discovery of DNA polymerase In mid-1950s ARTHUR KORNBERG showed that an enzyme isolated from bacterial cells can carry out DNA synthesis in a test tube This enzyme is called DNA polymerase

Discovery of Polymerase II & III:

Discovery of Polymerase II & III In 1969 PETER DELUCIA AND JOHN and is associate report that mutant stain of bacteria defective in kornberg enzyme It can replicate the DNA and divided in normal manner without kornberg enzyme

Slide 18:

They detect presence of two other bacterial enzymes the synthesis DNA This enzyme are named DNA polymerase II & III

Slide 19:

Property Pol I Pol II Pol III (core)* Mass(kd) 103 90 130, 27.5, 8.6 Turnover No + 600 30 1200 Structural gene PolA Pol B dnaE (a subunit) dnaQ (e subunit) holE (q subunit) Polymerization 5' ® 3' Yes Yes Yes Exonuclease 5' ® 3' Yes No No

Slide 20:

DNA polymerase contain member of site which have functional importance 1-Template site 2-Primer site 3-Triphosphate site A triphosphate site where incomeing nucleoside triphosphate matches a complementary to the 3’OH position of primer

Polymersization 5’-3’ end :

Polymersization 5’-3’ end The synthesis of new DNA chain from its nucleotides is called polymerization It polymerase at rate of about thousand molecule per minute It also perform clean up function during replication recombination and repair

Slide 23:

The special function are enhance by its 5’-3’exonuclease activity when the 5’-3’exonuclease domain is removed the remaining fragment called the large or KLENOW fragments which retain polymerization

Polymerase II:

Polymerase II DNA polymerase II show significant activity only with gaped DNA template DNA synthesis take place in 5’-3’ direction it also require 3’OH primer Poly – II an elongate okazai fragment in the absence of polymerase I

POLYMERASE III:

POLYMERASE III Polymerase III exits in a complex holoenzyme form in the cell It as both 3’-5’ and 5’-3’ exonuclease DNA poly – III cannot be used single strand DNA molecule act as a template even provide with a primer

Slide 26:

Subunit Mass kD Structural Gene Function Alpha 130 polC(dnaE) Polymerase epsilon 27.5 dnaQ 3 exonuclease 8.6 holE a, assembly 71 dnaX Assembly of holoenzyme on DNA Beta 41 dnaN Sliding clamp, processivity Gamma 47.5 dnaX(Z) Part of the gamma complex* Delta 39 holA Part of the gamma complex* Delta’ 37 holB Part of the gamma complex* 17 holC Part of the gamma complex* 15 holD Part of the gamma complex* The Subunits of E. coli DNA Polymerase III Holoenzyme

Proteins Involved in DNA Replication :

Proteins Involved in DNA Replication Protein Function DNA gyrase Unwinding DNA SSB Single-stranded DNA binding DnaA Initiation factor HU Histone-like (DNA binding) PriA Primosome assembly, 3' ® 5' helicase PriB Primosome assembly PriC Primosome assembly DnaB 5' ® 3' helicase (DNA unwinding) Primase Synthesis of RNA primer DNA polymerase III holoenzyme Elongation (DNA synthesis) DNA polymerase I Excises RNA primer, fills in with DNA DNA ligase Covalently links Okazaki fragments

In Eukaryatic DNA:

In Eukaryatic DNA DNA polymerase ALPHA DNA polymerase BETA DNA polymerase gamma DNA polymerase delta DNA polymerase epsilon

DNA polymerase ALPHA:

DNA polymerase ALPHA Is reponsible for synthesis of RNA primer for both leading and lagging strands DNA polymerase BETA Is involved in their repair of DNA it function is comparable with DNA polymerase I in prokaryotes

Biochemical properties of eukaryotic DNA:

Biochemical properties of eukaryotic DNA Mass(KD) Alpha Delta Epsilon Beta Gamma Native >250 170 256 36-38 160-300 Catalytic core 160-180 125 215 36-38 125 Other subunit 70,50,60 48 55 None 35,47 Location Nucleus Nucleus Nucleus Nucleus Mitochondria Associated function 3’-5’exonuclease No Yes Yes No yes Properties Processivity Low High High Low High Fidelity High High High Low High Replication Yes Yes Yes No Yes Repair No ? Yes Yes no

Slide 31:

DNA POLYMERASE GAMMA These enzyme participate in their replication of mitochondrial DNA DNA POLYMERASE DELTA Is responsible for the replication on the leading strand of DNA it is also possess proof reading activity DNA POLYMERASE EPISLON is responsible for the replication on the lagging strand of DNA it is also possess proof reading activity

Slide 32:

Thank you Meet again