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Genetics & Evolutionary Biology > DNA Repair Mechanisms > Flashcards

DNA Repair Mechanisms Flashcards

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1
Q

What is a Thymine Dimer?

A
  • A pair of abnormally chemically bonded adjacent thymine bases in DNA, resulting from damage by ultra-violet irradiation
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2
Q

How does Light dependent Repair work?

A
  • A Photolyase Cleaves the Thymine Dimers
  • Photoreactivation performed by the light activated enzyme DNA photolyase
  • Step 1: DNA exposed to UV-> T dimers
  • Step 2: DNA photolyase binds T dimers
  • Step 3: DNA photolyase activated by blue light -> cleaves T dimers
  • Step 4: DNA photolyase released
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3
Q

What steps are involved in Excision repair?

A
  • A DNA repair endonuclease or endonuclease-containing complex recognizes, binds to, and excised the damaged base or bases
  • A DNA Polymerase fills in the gap, using the undamaged complementary strand of DNA as a template
  • DNA ligase seals the break left by DNA polymerase
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4
Q

What are the types of Excision Repair?

A
  • “Base excision repair” pathways remove abnormal or chemically modified bases
  • “Nucleotide excision repair” pathways remove larger defects, such as thymine dimers
  • Both occur by similar mechanism in E. coli & Humans
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5
Q

What are the first three steps of Base Excision Repair?

A
  • Inititated by DNA glycosylases
  • Step 1: Modified base
  • Step 2: Specific glycosylase binds specific altered based
  • Step 3: Specific glycosylase cleaves the glycosidic bond between the abnormal base and 2- deoxyribose, creating apurinic or apyrimidinic sites (AP sites) with a missing base
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6
Q

What are the 4th-6th steps of Base Excision Repair?

A
  • Step 4: AP site recognized by AP endonucleases that act together with phosphodiesterases to excise the sugar-phosphate groups at this site
  • Step 5: DNA polymerase replaces the missing nucleotide according to the specifications of the complementary strand
  • Step 6: DNA ligase seals the nick
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7
Q

What three genes are required for Excinuclease activity?

A
  • Excinuclease activity requires the products of three genes, uvrA, uvrB, & uvrC
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8
Q

What are the first 2 steps in Nucleotide Excision Repair?

A
  • Step 1: A trimeric protein containing two UvrA and one UvrB recognizes and binds the defective DNA
  • Step 2: Energy from ATP used to bend the DNA at the damaged site. The UvrA dimer is then released
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9
Q

What are the 3rd and 4th steps of Nucleotide Excision Repair?

A
  • Step 3: UvrC binds to the UvrB/DNA complex and UvrB cleaves the 5th phosphodiester bond from the damaged nucleotide(s) on the 3’ side, and the UvrC hydrolyzes the 8th phosphodiester linkage from the damage on the 5’ side
  • Step 4: UvrD (DNA helicase II) releases the excised dodecamer
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10
Q

What are the 5th and 6th steps of Nucleotide Excision repair?

A
  • Step 5: DNA polymerase I replaces UvrB protein and fills in the gap using the complimentary strand as a template.
  • Step 6: DNA ligase seals the nick left by polymerase
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11
Q

What is the difference between Nucleotide Excision Repair in humans and in E. coli?

A
  • Nucleotide excision occurs by a similar pathway in humans, except that many more proteins are involved and a 24- to-32 nucleotide long oligomer is excised
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12
Q

What is Postreplication Mismatch Repair?

A
  • Provides a backup to the replicative proofreading activity of DNA polymerase by correcting mismatched nucleotides remaining in DNA after replication
  • System must be able to distinguish between the template (parental) strand and the newly synthesised strand (that contains the mismatched base)
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13
Q

What is Methylation?

A
  • DNA methylation is a process by which methyl groups are added to the DNA molecule.
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14
Q

In Postreplication Mismatch Repair in E. coli, what is Methylated?

A
  • The A in GATC sequences is methylated subsequent to DNA replication
  • In newly replicated DNA, the parental strand is methylated, but the new strand is for a time not. This difference allows the mismatch repair system to distinguish the new strand from the old strand
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15
Q

What is excised in Postreplication Mismatch Repair in E. coli

A
  • The mismatched nucleotide is excised from the new strand and replaced with the correct nucleotide, using the methylated parental strand as a template
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16
Q

What does Postreplication Mismatch Repair in E. coli require?

A
  • Requires the products of four genes:

mutH, mutL, mutS, and mutU

17
Q

What are the four genes for postreplication mismatch repair in E. coli used for?

A
  • MutS recognizes mismatches and binds to them to initiate the repair process
  • MutH and MutL join the complex
  • MutH cleaves the unmethylated strand at hemimethylated GATC sequences on either side of the mismatch
  • Excision requires MutS, MutL, MutU (DNA helicase II), and an exonuclease
  • DNA polymerase III fills in the gap, and DNA ligase seals the nick
18
Q

Study the diagrams for Light Dependent repair, The Base Excision Repair Steps and Postreplication Mismatch Repair in E. coli

A

https://docs.google.com/document/d/1Nzo4FTzXCbwOZjpoc_J_4IF3gsOXPcoyC2BowELmx0U/edit?usp=sharing

19
Q

How is the SOS Response in E. coli activated?

A
  • If DNA is heavily damaged by mutagenic agents, the SOS response, which involves many DNA recombination, DNA repair, and DNA replication proteins, is activated
20
Q

How does the SOS Response in E.coli work?

A
  • DNA Polymerase V replicates DNA in damaged regions, but sequences in damaged regions cannot be replicated accurately
  • This error-prone system eliminates gaps but increases the frequency of replication errors
21
Q

How is the SOS Response cancelled when there is no damage of the DNA?

A
  • In the absence of DNA damage, LexA binds to DNA regions that regulate transcription of SOS response genes and keeps their expression levels low
22
Q

What happens when the SOS Response is needed but the LexA has binded to the DNA?

A
  • When extensive DNA damage occurs, RecA binds to single-stranded regions of DNA in damaged regions
  • This activates RecA, which stimulates LexA to inactivate itself. When LexA is inactivated, the SOS response genes are expressed
23
Q

How many human disorders are there from DNA Mutations?

A
  • Several inherited human disorders result from defects in DNA repair pathways
  • For example: Xeroderma Pigmentosum (XP)
  • Individuals with XP are sensitive to sunlight
  • The cells of individuals with XP are deficient in the repair of UV-induced damage to DNA
  • Individuals with XP may develop skin cancer or neurological abnormalities
24
Q

What does DNA Recombination involve?

A
  • Recombination between homologous DNA molecules involves the activity of numerous enzymes that cleave, unwind, stimulate single-strand invasions of double helices, repair, and join strands of DNA
25
Q

How does Recombination work?

A
  • In eukaryotes, crossing over is associated with the formation of the synaptonemal complex during prophase of meiosis I
  • Crossing over involves the breakage of parental chromosomes and rejoining of the parts in new combinations
26
Q

Study the diagrams for SOS Response, DNA Recombination and Crossing over

A

https://docs.google.com/document/d/1Nzo4FTzXCbwOZjpoc_J_4IF3gsOXPcoyC2BowELmx0U/edit?usp=sharing

27
Q

What models are used in the molecular basis of Recombination?

A
  • The Holliday model and the double-strand break model are two explanations of the molecular basis of recombination
28
Q

What are the first 3 steps in the Holliday model?

A
  • Endonuclease cleaves single strand of each parental DNA molecule
  • Segments of the single strand on one side of each cut displaced from their complementary strand by action of helicase
  • Helicase unwinds the two strands of DNA in the region adjacent to single-strand incision
29
Q

What are the 4th and 5th steps in the Holliday model?

A
  • Stimulated by RecA-type proteins, the displaced single strands exchange pairing partners, base pairing with the intact complementary strands of the homologous chromosomes
  • Cleaved strands are covalently joined by DNA ligase
30
Q

What are the 6th and 7th steps in the Holliday model?

A
  • An X-shaped recombination intermediate called chi structure results (has been observed by electron microscopy)
  • Endonuclease catalyses the cleavage of single strands at the chi junction
31
Q

What is the final step of the Holliday model?

A
  • Following cleavage at chi junction the complementary DNA strands are rejoined to produce two recombinant DNA molecules
32
Q

Study the diagrams for the Holliday model and for the 2 types of Recombination.

A

https://docs.google.com/document/d/1Nzo4FTzXCbwOZjpoc_J_4IF3gsOXPcoyC2BowELmx0U/edit?usp=sharing

Genetics & Evolutionary Biology (16 decks)

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