1C: DNA replication and repair Flashcards
what is the semiconservative model of DNA replication?
each daughter strand remains paired with its complementary parental strand (so each parent DNA transforms into two daughter DNA where one strand is new but one comes from the parental strand)
what is the conservative model of DNA replication?
both daughter strands pair up and both parent strands pair up.
what is the dispersive model of DNA replication?
daughter strands have a mixture of parental and newly synthesized DNA
what were the steps of Meselson and Stahl’s experiment?
1) bacteria grown in 15N (heavy nitrogen) medium. the isotope was incorporated into the nitrogenous bases of their DNA.
2) bacteria transferred to 14N (light nitrogen) medium, resulting in all newly formed nitrogenous bases being light.
3) DNA extracted from bacteria after each generation in new media. centrifuged in special solution to separate DNA of different densities
what were the results of the Meselson and Stahl’s experiment? how did this compare to the potential predicted results?
after one generation, all DNA was mid weight. after two generations, some DNA was mid weight and some was light.
this is aligns with the predicted weights for the semiconservative model of replication.
what were the predicted banding patterns for the 3 replication models?
➞ semiconservative: after gen 1 is all mid weight, after gen 2 is some mid weight some light weight.
➞ conservative: after gen 1 and after gen 2 both have some heavy and some light.
➞ dispersed: after gen 1 is mid weight, after gen 2 is mid but slightly lighter
to which end can nucleotides be added?
only the 3’-OH end
in what direction does DNA synthesis occur?
5’ to 3’
what provides energy for formation of new phosphodiester bond?
hydrolysis of pyrophosphate
what are 2 properties of polymerases?
1) cannot synthesize new strands from scratch (must build off a primer)
2) has a single active side that can catalyze 4 different reactions (incorporation of the 4 DNA bases)
what is a replisome?
molecular machine made up of all the enzymes that replicate DNA
what does helicase do?
unwinds the double helix by breaking hydrogen bonds
what does primase do?
synthesizes RNA primers for DNA polymerase
what does single-strand binding protein do?
stabalizes ssDNA (single-stranded) before replication by preventing reannealing so the strands can serve as templates
what does topoisomerase do? (what is an alternate name for it?)
removes twistiness of DNA ahead of the replication fork by cleaving the DNA, turning it, and reattaching
(alt name: gyrase)
what does polymerase III do?
synthesizes DNA by adding nucleotides to primer
what does polymerase I do?
removes the RNA primer and fills the gaps left by the primer with DNA
what does the sliding clamp do?
attaches polymerase III to the DNA template. makes replication more efficient (smoother, faster)
what does ligase do?
joins the ends of the DNA segments by forming phosphodiester bonds
what are the two new strands of DNA formed by the replication fork called?
➞ leading strand
➞ lagging strand
what is the leading strand?
the strand that is synthesized continuously (synthesized in the same direction as the movement of the replication fork)
what is the lagging strand?
the strand that is discontinuously synthesized (synthesis is in the direction opposite to the movement of the replication fork)
what are Okazaki fragments?
small DNA fragments formed during synthesis of the lagging strand
in what direction is the template strand read in DNA replication? why?
3’ to 5’ because it must be build 5’ to 3’ and the strands must be antiparallel
what is initiation? (DNA replication of bacterial chromosomes)
unwinding and separation of the two template DNA strands at the origin of the replication site (ori)
what is elongation? (DNA replication of bacterial chromosomes)
simultaneous synthesis of the two DNA strands from the template strands by DNA polymerase
what is termination? (DNA replication of bacterial chromosomes)
DNA replication stops upon reaching termination site or the end of the chromosome
what is the end replication problem?
since RNA primers are required for polymerases to build off of, the RNA that connects to the very beginning of the strand has no way of being replaced by DNA, which would result in the DNA strand being shortened with every replication.
what is the solution to the end replication problem?
the telomere (which is grown in gametes and stem cells)
what is telomerase?
enzyme that restores shortened telomeres.
how does telomerase work?
it connects to the 3’ end of the DNA with some RNA on the bottom (as a template). it builds more DNA onto the 3’ end of the template DNA using the telomerase RNA as a template. then telomerase disconnects and some primer puts down RNA and starts the normal DNA replication cycle over again
where is telomerase present?
gametes and stem cells (this is why newborns have longer telomeres than their parents).
NOT present in most eukaryotic cells, so telomeres shorten as an individual ages
how can cancers use telomerase?
many cancers acquire mutations that activate telomerase to negate the limitation of rapid cell division, resulting in formation of tumours that don’t stop growing on their own like normal cells.
what is a telomere?
a buffer of highly repetitive noncoding DNA that protects actual genes from the shortening that occurs with every cell division
what are the 3 types of damage repair mechanism that correct DNA damage?
➞ proofreading
➞ mismatch repair
➞ excision repair
what is the process of proofreading?
1) polymerase III is synthesizing the new strand
2) polymerase (rarely, 1/10 000 times) adds incorrect base
3) polymerase recognizes the mistake, reverses, and uses exonuclease (which moves 3’ to 5’) to remove the nucleotide from the strand
4) polymerase resumes polymerization
what is the chance of mismatches caused by polymerase (including proofreading)?
1/10 000 000
what is the process of mismatch repair (MMR)?
1) binding protein MutS and MutL recognize mismatch damage.
2) MutH endonuclease nicks daughter strand sever nucleotides away from the mismatch
3) exonuclease Exo1 excises region of daughter strand surrounding the mismatch in the 5’-3’ direction
4) polymerase III fills gap and repairs mismatch
5) nick left after gap sealed by ligase
which errors does MMR deal with?
replication errors not corrected by proofreading
