R6 DNA replication Flashcards
What does each carbon in ribose interact with ?
1’ - base
2’ - OH in RNA, no OH in DNA
3’ and 5’ - phosphate
What is a nick and a gap ?
A nick is one strand of the sugar-phosphate is broken, no missing nucleotides, the phosphodiester bond is broken between the nucleotides, needs to be sealed by ligase
A gap is a missing stretch of the nucleotide, usually occurs on the lagging strand. Needs to be filled in by DNA polymerase and then sealed by ligase
Where do the arrows run in a diagram of replication ?
5’ to 3’
What are the 3 types of ends on double stranded DNA ?
Blunt end, 3’ overhand, 5’ overhang
What do nuclease, ligase and primase do ?
Nuclease – cleaves sugar-phosphate backbone
Ligase – joins sugar-phosphate backbone
Primase - synthesis of primers
what type of junction does a replication fork show ? a repair intermediate ?
replication fork - 3 way
repair intermediate - 4 way
What are the types of replicative helicases in prokaryotes and eukaryotes ?
DnaB is the replicative helicase (5’3’) in prokaryotes, MCM is the replicative helicase (3’5’) in eukaryotes.
Why are helicases called molecular motors ?
They use ATP, they turn energy into movement
re-annealing afterwards does not require energy
What form do free nucleotides come in ?
Triphosphate, so when it is de-phosphorylated it releases energy for replication
What are the 3 types of replication ?
Conservative vs semi-conservative vs dispersive
What does an exonuclease do ?
corrects the wrong base and puts the right one in
What does topioisomerase do ?
works further upstream to prevent mechanical stress
DNA synthesis can only happen in … to …
5’ , 3’
What occurs on the lagging strand ?
- RNA primers are laid on the lagging strand, and synthesized by primase
- DNA pol I then removes the primers and fills the gap
- DNA ligase fills in the gaps
What do single-strand binding proteins (SSB) do ?
Bing to ssDNA to prevent winding back
What do sliding clamps do ?
hold the polymerase in place whilst nucleotides are being added
What does DNA pol I do ?
removes RNA primers and replaces them with DNA
What is the differences in the replication fork in eukaryotes to prokaryotes ?
leading and lagging strand is synthesized by different DNA polymerases
Okazaki fragments synthesis
the replicative helicase travels along the lagging strand in prokaryotes and the leading strand in eukaryotes
What are telomeres ?
the ends of linear eukaryotic chromosomes. Highly repetitive G-C and prevent degradation
What are shelterin proteins ?
protect the telomeres and cap the chromosomes as a whole
What happens to telomeres with every replication cycle ?
the telomeres shorten due to the leading strand being shorter than the lagging due to the lack of ozahaki fragments
The shorter the telomeres the more limited cell proliferation is
The shortening of telomeres has a protective role against cancers, they limit proliferation of pre-cancer cells.
Cancer formation consists of stabilising the telomere length
What can nuclease do ? What are the different types of nucleases ?
break the DNA sugar-phosphate backbone between the phosphate and the C3 or between the phosphate and the C5
There are ssDNA specific nucleases and dsDNA specific nucleases
Exonucleases begin cleaving at the ends (requires ends to the linear DNA) of the DNA (5’ and 3’ or 3’ and 5’), endonucleases break mid-strand (don’t require an ending)
How can helicase and nuclease couple ?
Helicase can bind (as a substrate) with the nuclease, to degrade both or one strand. If one strand degrades then the 3’ overhang allows for a template for DNA repair.
What do replicase, reverse transcriptase, transcriptasem RNA replicase do ?
Replicase – polymerases for DNA replication
Reverse transcriptase – telomerase and the ones coded by retrovirus
Transcriptase – RNA polymerases in prokaryotes and eukaryotes, primase
RNA replicase – RNA viruses other than retrovirus undergo RNA replication and code for RNA replicase
What is the directionality of polymerase ?
All polymerase have 5’ 3’ directionality, so you cannot extend the 5’ end, you can only degrade it
What is the proofreading activity of the DNA polymerase ?
-DNA polymerases has a proofreading activity, removes a wrong base, which is done by a 3’–> 5’ exonuclease
Proofreading activity can be removed, if mutation is needing to be high (to create mutants)
Processivity is the ability of the polymerase to extend a DNA strand before it dissociates from the DNA and needs to bind again, the affinity for DNA aka.
What does nascent mean ?
made or derived from
What is a clump ?
a closed ring which cannot fall off the DNA, it slides along the DNA
Most common is PCNA (proliferative cell nuclear antigen), can react with 3 proteins at a time, brings different enzymes to the replication forms, it is regulated by SUMOylation and ubiquitination
How does clump loading occur ?
Clump loader binds to the DNA and then the clump, then the loader is released.
Clumps do not catalyse
Clump loader is an enzyme as it catalyses the opening and closing of the clump ring using ATP.
