DNA replication Flashcards
What are the requirements for DNA replication to occur?
1) a single stranded template
2) deoxyribonucleotide triphosphate (dNTPs) of (A,G,C,T and Mg2+)
3) replisome: a nucleoprotein complex that co-ordinates the replication actives numerous enzymes and proteins
4) a primer with a free 3’ end hydroxyl group
where does the replication process begin on the DNA sequence?
- separation of two complimentary strands occurs at the ‘origins of replication’
What are the ‘unwinding proteins’ ?
- DNA helicase - separates the DNA strands in an ATP dependent process
- single-stranded binding proteins bind to prevent the strands from re-associating
- Topoisomerase - regulates the twisting of the DNA - by cutting the DNA- ligase then comes and reanneals the two together
What enzymes are used to replicate/copy the DNA in replication?
- primase - makes a primer and starts a daughter strand
- DNA polymerase
Why do we need ‘priming’ sequences for DNA replication to occur?
because DNA replication requires a free 3’ hydroxyl group to extend a DNA chain on- so in order to go in both directions of the DNA strands, we require primers
In what direction does DNA polymerase read it’s template?
3’ to 5’
In what direction does DNA polymerase make a new DNA chain?
5’ to 3’
DNA polymerase catalyses the formation of what bonds?
It catalyses the formation of phosphodiester bonds
What is the role of the PCNA (proliferating cell nuclear antigen)?
it acts as a ‘clamp’ associated with DNA polymerase, that essentially keeps the DNA pol. bound to the template as it rapidly reads the template and produces the DNA strand
How do wee check errors in cell replication?
1) substrate specificity - the DNA pol active site can bind all four dNTP types, but catalysis only occurs when the correct one is bound
2) proof-reading- error correction activates 3’ to 5’ exonuclease activity (in reverse direction) it removes nucleotides at the 3’ end of a new strand that are mismatched
Describe the differences in activity between
DNA polymerase epsilon, delta, beta, and gamma
epsilon = leading strand synthesis
delta = lagging strand synthesis
beta= involved in DNA repair
gamma = replicates mitochondrial DNA
what is the purpose of telomeres in DNA replication?
Because daughter DNA must always add to an already existing 3’ hydroxyl group, there will always be a small loss ( where the primer sat) where DNA cannot form. This is the purpose of telomeres- they are non-coding portions.
What is a ‘mismatch repair’?
occurs shortly after replication, replaces mismatched bases or loops up to 4 bas pairs long in DNA -
How to repair systems know which strand is the parent vs. daughter strand?
methylation is on the parent strand, the daughter strand is not methylated
What is ‘base excision repair’?
it replaces bases lost through chemical processes (depurination or deamination)
- DNA glycosylase identifies and removes damaged bases
- AP endonuclease cuts the backbone
- Exonuclease removes the sugar and several adjacent bases
