Lecture 3: DNA Replication, Repair and Recombination 1 Flashcards
- Review DNA replication - Learn pathways for DNA repair
Why do multicellular organisms need high fidelity?
- germ cells need it to have low mutation rates to maintain the species
- somatic cells ned low mutation rates to avoid uncontrolled proliferation/cancer
What are the requirements of DNA replication?
- separation of the two parental strands
- dATP, dGTP, dCTP, and dTTP
- DNA polymerase requires a primer with a free 3’ OH to begin
When does exonucleolytic proofreading occur?
immediately after incorrect base is added
What happens during exonucleolytic proofreading?
- DNA polymerase requires a perfectly paired 3’ terminus
- 3’ to 5’ exonuclease clips off unpaired residues at 3’ primer terminus
What is DNA helicase?
- unwinds DNA
- 6 identical subunits that bind and hydrolyze ATP
What are single stranded binding proteins?
-bind highly and cooperatively to exposed SS DNA
What does single stranded binding proteins do?
- help stabilize unwound DNA
- prevent formation of hairpins
- DNA bases remain exposed
What does the sliding clamp do?
Keeps DNA polymerase on DNA when moving, releases when double stranded DNA is encountered
Assembly requires a _______ which _______
clamp loader; hydrolyzes ATP as it load the clamp onto a primer template junction
What does the clamp do with the leading strand of DNA?
clamp remains associated to DNA polymerase for long stretches
What does the clamp do with the lagging strand?
clamp loader stays close so it can assemble a new clamp at start of each Okazaki fragment
What does mismatch repair do?
removes (almost all) errors missed by proofreading by detecting distortion caused by mispairing
What is DNA topoisomerase?
reversible enzyme that breaks a phosphodiester bond to change superhelicity, relieving supercoiling
How does type I topoisomerase work?
creates single strand break in DNA which allows the DNA on either side of the nick to rotate freely relative to each other; uses phosphodiester bond as a swivel point
Which type of topoisomerase is thermodynamically favorable?
Type I
How does type II topoisomerase work?
Makes a transient double strand break. Uses ATP to:
- break one double stranded reversibly to create gate
- causes second stand to pass through
- reseals break and dissociates
What is a replication origin rich in?
A-T sequences
What does it mean by the refractory period referring to replication origins?
delay until new strand in methylated
Minimum requirements for sequence to be ORI
- binding site for origin recognition complex (ORC)
- A-T rich stretch for easy unwinding
- binding site fro proteins that help attract ORC
What is required for chromatin reassembly after replication?
histone chaperones
What sequence is at the end of chromosomes and repeated?
GGGTTA
What do telomerase do?
replenish end sequences by elongating parenting stand in 5’ to 3’ direction using an RNA template on the enzyme
____ cells retain full telomerase activity.
stem
What is replicative senescence?
when daughter cells have defective chromosomes and stop dividing; in this way the cell’s lifetime is regulated to guard against cancer
