The rolling circle replication model forms what?
One circular molecule and one linear molecule, which may circularize.
Explain the steps of prokaryotic replication
- DNA A proteins bind to Ori C site.
- DNA A and C recruit DNA helicase (DNA B), and DNA C is the clamp loader that loads DNA B onto the strand, which starts unwinding the DNA.
- SSBP help to keep the strands from reannealing, making replication bubble
- (DNA G) primase places an RNA primer on both strands
- DNA pol III comes in, and the CLAMP LOADER (gamma subunit of pol III) helps load it onto template strand. Pol III beta subunit is sliding clamp
- DNA pol III proceeds w/replication
- RNA primers are replaced with DNA (dNTP) by DNA pol I (Nick translation) and RNaseH
- DNA ligase connects the DNA to the rest of the strand
- Tus protein terminates it by binding to ter sequence, stalling helicase, resulting in catenation (also dimerizes core complex)
- DNA gyrase/Topo II decatenate
What prokaryotic DNA polymerase has 5’-3’ exonuclease activity? Why is this important?
DNA polymerase I. This allows it to remove primers!!
DNA gyrase is what type of topoisomerase? what type of strand does it work on? how many breaks does it make?
Topo II. Works on negatively coiled strands. Double-stranded break.
What does acyclovir do? Used to treat what?
Stops RNA pol function. Used to treat herpes.
What does AZT (azidothymidine) do? Used to treat what?
Analog of thymine base and a chain terminator, used to treat HIV.
dideoxyinosine used to treat what?
AIDS.
antimalarial drugs target what?
target both polymerase and topo II
Nalidixic acid and norfloxacin do what? Treatment for?
Target bacterial DNA gyrase to inhibit replication. Used to treat UTIs.
explain eukaryotic replication
- Pre Replication complex makes origin. ORC+cdt1 bind to DNA
- Recruits and forms a Helicase called CMG complex, made of Cdc45, MCM2-7 and GINS.
- RPA acts a ssbp to keep strands apart.
- Pol alpha/primase adds RNA primer to DNA strands.
- RFC (clamp loader) loads PCNA (DNA clamp) onto DNA
- Polymerase epsilon synthesizes the DNA in leading strand, and pol delta does okazaki fragments.
- RnaseH+Fen1+Pol-delta remove and replace RNA primers
- DNA ligase seals it all.
Do eukaryotic polymerases have 5’-3’ exonuclease activity?
NO, but do have 3-5
Explain telomerase action (euk, obviously)
- Telomere is a ribonucleoprotein that is a reverse transcriptase (TERT)
- telomerase has built-in RNA template (TERC) complementary to the G rich overhang
- Telomerase adds DNA telomere repeats, has reverse transcriptase
- Telomerase leaves, and polymerase fills in complimentary strand.
- nucleosomes reform, epigenetic pattern copied
What do CAFs do (chromatin assembly factors)?
They help in making new histones, as both helices need the histone proteins.
What makes DNA pol delta processive in euk?
What about for DNA Pol III in Prok?
(these are sliding clamps)
PCNA
Beta-subunit of Pol III
SSBP and RPA have similar functions. which is for prok and which is for euk?
SSB for prok, RPA for euk
Prokaryotic replication: How many origins? Replication model? uni or bidirectional? Shape of strand? continuity? breakage of strand? products?
One origin Theta model Bidirectional circular DNA semidiscontinuous no break in strand two circular
viral replication: Uni or bidirectional? replication model? breakage of strand? product?
unidirectional
rolling circle
yes, breaks strand
one circular and one linear product
which prok polymerase has 3’-5’ exonuclease activity?
Pol III, II, I
unlike prokaryotes, eukaryotes need NO helicase loader!
1
what is the fork protection complex in euk?
links helicase and polymerase.
both prok and euk polymerase have 3-5 exo, but only prok has 5-3.
.
helicase for prok and for euk?
Prok- DNA B
Euk- CMG
primase prok and euk?
prok- DNA G
euk- polymerase alpha primase
RNA primer replacement prok vs. euk
prok- RNaseH and Pol I
euk- RNaseH1 and Fen1 and Pol delta
