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Translession DNA Synthesis

  1. site specific recombination is illegitimate.. Transposase enzyme recognized the self DNA and cut it and also recognized the target site. Target site is not specific .
  2. Transposons
  3.  Reterovirus Like elements (RLE)- like reterovirus  42 % human genome made up on reterotransposons ..
  4. Repair system Direct Repair 1) Photoreactivation Excision Repair BER NER Mismatch Transcriptional coupled repair system Double stranded Break Repair 1.Homologous recombination 2. Non-homologous End joining Mech: Single stranded DNA Repair (Translession DNA Synthesis) SOS heavy damage  translesion polymerases Any type of damage remove before cell division Genetic information can be stored stably in DNA sequences only because a large set of DNA repair enzymes continuously scan the DNA and replace any damaged nucleotides.
  5. • Translession DNA Synthesis • oxidizing agents , metabolites , radiation and reactive chemicals DNA suffers heavy damage  translesion polymerases • When DNA damage is excessive, a special class of inaccurate DNA polymerases, called translesion polymerases, is used to bypass the damage, allowing the cell to survive but sometimes creating permanent mutations at the sites of damage. • Y- Family DNA polymerase • Lack of proofreading Activity • Add nucleotide at lesion without base pairing DNA Polymerase IV ---Din B Gene DNA Polymerase V – UMU, UMUD Gene /------ Repressor Lex -A SOS Repair / Translession DNA Synthesis / Error prone Repair
  6. DNA Polymerase IV ---Din B Gene DNA Polymerase V – UMU, UMUD Gene /------ Repressor Lex –A In normal condition Lex –A Repressor protein bind to Din B , UMU, UMUD Gene . And inhibit expression of translesion polymerases. Excessive DNA Damage or single strand DNA Damage = High Rec-A Protein Active and Interact with Lex-A and Auto-Protease Activity of Lex-A is on. Cleaved Lex-A – not bind to Din B , UMU, UMUD Gene DNA Pol- IV DNA Pol-V Absent Proofreading Activity Add nucleotide at lesion without base pairing
  7. Double strand break repair • Prokaryotic and lower eukaryotic – Homologous Recombination • Higher eukaryotic – Non – Homologous End Joining (NHEJ) Error prone Repair mechanism .
  8. HOMOLOGOUS RECOMBINATION • the mechanisms that allow the DNA sequences in cells to be maintained from generation to generation with very little change. • repair mechanism is essential for every proliferating cell • homologous recombination (also known as general recombination)is an exchange of DNA strands between a pair of homologous duplex DNA • Work in meiosis in plants and animals. • daughter DNA molecules are still held close together. homologous recombination  flexible series of reactions
  9. Prokaryotics RecA first binds cooperatively to the invading single strand, help to pairing of homologous strands. Rec B,C,D – processing of ds DNA break. 5’3’ exonuclease 3’5’ Exonuclease strand exchange protein complex DNA Pol III and DNA Ligase DNA synthesis and ligation RUV-A,B – Branch Migration steps (Helicase Activity) RUV-C – Resolution ( Holiday Junction)  Both strand separate.
  10. Homologous recombination in eukaryotic • DNA Damage / Mutant sensor Kinase – ATM (Ataxia-Telangiectasia Mutated kinase) . • Sensory Kinase- chk-2 . • Pairing of Homologous strands – BRCA-1, BRCA-2, WRN, Rad-5 . • Processing of double strand Break – Rad-50,Rad 58,Rad 60 or MRX complex in Yeast. • Strand exchange protein Assembly – Rad 52, Rad 58. • Resolution ( Holiday Junction)  Both strand separate  Nuclease or resolvase
  11. NHEJ If these lesions were left unrepaired , they would quickly lead to the breakdown of chromosomes into smaller fragments and to loss of genes . Nonhomologous end joining  loss of nucleotides at the site of joining . end-joining mechanism “quick and dirty” solution to the repair of double-strand breaks  common in mammalian somatic cells . Age of 70. Nonhomologous end joining  one broken chromosome becomes covalently attached to another. This can result in chromosomes with two centromeres and chromosomes lacking centromeres.
  12. NHEJ 1. Recognition  Ku protein 70/80  heterodimer binds to broken chromosome ends. 2. End processing-> Additional proteins (ARTEMIS,MRN) needed to hold the broken ends together while they are processed and eventually joined covalently. 3. Strand invasion , DNA synthesis and Resolution 4. Ligation- Lig-IV