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DNA Repair. Dr Derakhshandeh. For DNA. information must be transmitted intact to daughter cells. Accuracy is maintained by:. 1- High fidelity in replication 3’- exonuclease activity of DNA pol I Uracil-DNA N-glycosylase pathway (corrects mutations from deamination of

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Dna repair

DNA Repair

Dr Derakhshandeh


For dna
For DNA

  • information must be transmitted intact to daughter cells


Accuracy is maintained by
Accuracy is maintained by:

1- High fidelity in replication

  • 3’- exonuclease activity of DNA pol I

  • Uracil-DNA N-glycosylase pathway

    (corrects mutations from deamination of

    cytosine) cytosine Uracil


Accuracy is maintained by1
Accuracy is maintained by:

2-Mechanisms for correcting genetic info. in damaged DNA

  • e.gdue to chemical modifications

  • Irradiation changes


Categories of dna repair
Categories of DNA Repair

  • Mismatch Repair(Synthesis + Repairing)

  • MM created by replication errors

  • DNA Pol III proof reading

  • non-homologous recombination are recognized and corrected

DNA Pol III


Categories of dna repair1
Categories of DNA Repair

2. Base Excision Repair (Euk/Pro)

  • Starts at cleavage of glycosidic bond (connects base to sugar-phosphate backbone)

glycosidic bond


Categories of dna repair2
Categories of DNA Repair

3. Direct Repair- Damaged

base undergoes a chemical/UV

reaction Restores original

structure (pro)

  • e.g. DNA photolyase - E.coli


Categories of dna repair3
Categories of DNA Repair

4. Nucleotide Excision Repair (Prok: 12/Euk: 28)

- damaged DNA:

  • excised

  • replaced with normal DNA

    5. Recombinational Repair

  • Fills gaps in DNA :

  • Newly replicated DNA duplexes undergo genetic recombination

  • Removal of damaged segment


Dna repair1
DNA REPAIR

(1) Photoreactivation (aka Light Repair)


Direct dna damage and repair
DIRECT DNA DAMAGE AND REPAIR

  • A variety of irradiation (ionizing, ultraviolet, etc)

  • DNA damage of a variety of sorts:

  • U.V. induced formation of Thymine Dimmer

  • Blocked replication and gene expression until repaired

  • Prohotoreactivation enzyme

  • Photolyase

  • Prokaryote



T

C


Photoreactivation light repair
Photoreactivation (Light Repair)

  • PHR/PRE gene

  • codes for photolyase

  • with cofactor folic acid

  • binds in dark to T dimer

  • When light shines on cell

  • folic acid absorbs the light (photon)

  • uses the energy to break bond of T dimer

  • photolyase then falls off DNA


Dna repair2
DNA REPAIR

(2) Excision Repair

(aka Dark Repair)


Excision repair dark repair
Excision Repair (Dark Repair)

  • 3 different types of repair mechanisms

  • use different enzymes

  • (a) AP Repair (Base Excision Repair, BER)

  • (b) UV Damage Repair (also called NER - nucleotide excision repair)

  • (c) Mismatch Repair (MMR)


Ap repair base excision repair ber
AP Repair (Base Excision Repair, BER)

  • Repair of apurinic and apyrimidinic sites on DNA

  • in which base: has been removed

  • Base removed by:

    • DNA glycosylases

    • which remove damaged bases

  • ung gene codes for uracil-DNA glycosylase

    • recognizes and removes U in DNA

    • by cleaving the sugar-nitrogen bond to remove the base


AP endonucleases:

  • class I nick at 3' side of AP site

  • class II nick at 5' side of AP site

  • Exonuclease removes short region of DNA

  • DNA Pol I and ligase fill in gap


B uv damage repair also called ner nucleotide excision repair
(b) UV Damage Repair (also called NER - nucleotide excision repair)

  • It uses different enzymes

  • NER  removes a  large "patch" around the damage

  • Even though there may be only a single "bad" base to correct, its nucleotide is removed along with many other adjacent nucleotides

  • NER: UV

  • BER: Chemicals/Agents


Ner uv damage repair
NER (UV Damage Repair) repair)

  • Nuclease:

  • can detect T dimer

  • nicks DNA strand on 5' end of dimer (composed of subunits coded by uvrA, uvrB and uvrC genes)

  • UvrA protein and ATP bind to DNA at the distortion

  • UvrB binds to the UvrA-DNA complex and increases specificity of UvrA-ATP complex for irradiated DNA


  • UvrC repair) nicks DNA 8 bases upstream and 4 or 5 bases downstream of dimer

  • UvrD (DNA helicase II; same as DnaB) separates strands to release 12-bp segment

  • DNA polymerase I now fills in gap in 5'>3' direction

  • ligase seals

  • polA - encodes DNA pol I

    • mutant was viable retained normal 5'>3' exo activity

    • only 2% of polymerase activity



C mismatch repair mmr
(c) Mismatch Repair (MMR) repair)

  • Accounts for 99% of all repairs

  • Mismatch from replication

  • behind replication fork

  • Two ways to correct mistakes made during replication: 1) 3'>5' exonuclease - proofreading 2) Mismatch repair

  • mutL

  • mutS

  • mutH

  • andmutU(same UvrD) gene products involved (mut for mutator because if gene is mutated, cell has increased levels of spontaneous mutations)


How does system recognize progeny strand rather than parent strand as one with mismatch

Because of methylation repair)

DNA methylase (coded for by dam [DNA adenine methylase] locus)

methylates 5'-GATC-3' sequence in DNA at A residue

Mismatch from replication recognized by mutL and mutS gene products

mutH gene product nicks DNA strand (progeny strand) on either side of mismatch

DNA helicase II from mutUgene (also called uvrD gene)

unwinds DNA duplex and releases nicked region

Gap filled in by DNA Pol I and ligase

How does system recognize progeny strand rather than parent strand as one with mismatch?


Dna repair3
DNA REPAIR repair)

  • (1) Photoreactivation (aka Light Repair)

  • (2) Excision Repair (aka Dark Repair)

  • (3) Postreplicative (Recombinational) Translation Bypass Repair


Dna repair4
DNA REPAIR repair)

(3) Postreplicative (Recombinational) Translation Bypass Repair


Sos response
SOS response repair)

  • If T dimer is not repaired

  • DNA Pol III can't make complementary strand during replication

  • leaves large gap (800 bases)

  • Gap may be repaired by enzymes in recombination system

  • RecA - coats ssDNA

  • it also acts as autocatalysis of LexA repressor

  • recA mutants - very UV-sensitive

  • Now have sister-strand exchange - a type of recombination Translation bypass

  • Postreplicative repair is part of SOS response


Sos response1
SOS Response repair)


  • LexA repair) normally represses about 18 genes sulA and sulB, activated by SOS system

  • inhibit cell division in order to increase amount of time cell has to repair damage before replication

  • Each gene has SOS box in promoter

  • LexA binds SOS box to repress expression

  • RecA : LexA catalyses its own breakdown when RecA is stimulated by ssDNA

  • due to RecA binding ssDNA in lesions

  • could then bind to DNA Pol III complex passing through this area of the DNA and inhibit 3'>5' exonuclease (proofreading) ability

  • RecA no longer catalyzes cleavage of LexA (which is still being made)

  • so uncleaved LexA accumulates and turns the SOS system off


Why are dna repair systems necessary
Why are DNA Repair Systems Necessary? repair)

  • E.coli

  • Xeroderma Pigmentosum (XP)


E coli
E.coli repair)

  • repairing thymine dimers

  • important to bacteria

  • an E. coli strain that is:

    • phr (no photoreactivation)

    • recA (no translation by pass or SOS)

    • uvrA (no excision repair) is killed by a single thymine dimer


Xeroderma pigmentosum xp
Xeroderma Pigmentosum (XP) repair)

  • XP is a rare inherited disease of humans

  • predisposes the patient to:

    • pigmented lesions on areas of the skin exposed to the sun

    • an elevated incidence of skin cancer



  • It turns out that XP can be caused by mutations in any one of several  genes

  • all of which have roles to play in NER

  • Some of them:

  • XPA, which encodes a protein that binds the damaged site

  • assemble the other proteins needed for NER

  • XPB and XPD, which are part of TFIIH (Helicase)

  • XPF, which cuts the backbone on the 5' side of the damage

  • XPG, which cuts the backbone on the 3' side



Transcription coupled repair
Transcription-Coupled repair premature aging

  • Protein: ERCC6

    recognizes RNApol

    Mutation in gene:

    Cokayne Syndrom:

    MR

    Nerve disease

    Sensibility to sun


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