Chromosome biology lecture 9

Question 1

UV light + thymine dimers

Answer 1

• UV → photoproducts that are covalently linked btw adjacent pyrimidines (CPD)
• Issue as have covalent crosslinks that Δ ability to form bp + distort DNA ds helix

Question 2

NER

Answer 2

• Recognise damage, incision of affected DNA either side, excise, fill in w/ polymerase + ligase
• GC-NER vs TC-NER
• UvrA,B,C (mutagenesis)
• UvrA + B = important for proximal DNA damage recognition + act as platform
• UvrC = nuclease that acts on either side of damaged DNA, cuts 3’ to DNA
• UvrD = helicase that removes
• Eukaryotes = XPC + XPE, XPD (helicase that unwinds)
• In Tc-NER, recognition = DNA poly transcribing stalls, CSA + B associate = assembly point for factors

Question 3

NER + disease

Answer 3

• XP have defects in NER

Question 4

BER

Answer 4

• Similar to NER: recognise, remove, use undamaged strand as template

Question 5

DNA glycosylases

Answer 5

• Remove effected base by cleaving N glycosidic bond → apurinic site
• Carry out both recognition + excision
• Monofunctional = remove bases only leaving AP site
• Bifunctional = additional lyase that cleaves 3’ of basic site
• Recognise CHEMISTRY of base damage

Question 6

Uracil DNA glycosylase

Answer 6

• Spontaneous deam. of C→U = repaired
• UDGs are specific for U in DNA
• Scan along flipping out bases to see if fit in AS
• Specificity = bs too small for A + G, T has bulky C5 methyl so x fit

Question 7

AP endonuclease + DNA processing E

Answer 7

• Need 3’ to use as a primer
• AP cut leaving 3’OH
• E.g. Hunan apex1 has AP endonuclease, 3’ phosphatase and 3’ diesterase activity

Question 8

Different pathways

Answer 8

• Pathway II: DNA glycosylase cleaves N glycosidic bond → remove base → Apex1 cleaves 5’ site revealing 5’P + 3’OH, DNA pol fills in, ligase seals
• Pathway I: Type II glycosylase removes base → opening of ribose sugar + cleaving of backbone (x use for synthesis, need processing e.g. w/ Apex1)
• Pathway III: type II glycosylase give 3’P + 5’P, need to restore 3’OH (w/ PNKP) then DNA synthesis

Question 9

Oxidative damage

Answer 9

• ROS e.g. H202
• 2 main ways attack DNA:
• Addition to double bonds of bases OR H abstraction from deoxyribose sugars
• Radical attack → sugar fragmentation/base loss/ strand break

Question 10

DNA damage by ionising radiation

Answer 10

• IR directly damages through ionisation of base/sugar
• Due to H20 in system, species formed through radiolysis of H20 = main source of IR damage as → ROS
• 2 ss on opposite strands → DSB

Question 11

ss break repair signalling

Answer 11

• PARP = add ADP ribose onto proteins

- Secrete signals for repair proteins e.g. histones (opens structure)