(23)
5. Beskrive hvordan DnaA, DnaB+C, ssDNA og topoisomeraser er involveret i DNA
replikationen.
Devlin, s.169
Devlin, s.173 – fig.4.11
Devlin,
s.171 - fig.4.8
DnaA, DnaB and DnaC have an important role in initiating the replication process. (fx E.Coli)
DnaA – also called initiator protein. It binds to the origin of replication, where it recognizes 4 sequences of 9 nucleotides that are methylated. 4 DnaA molecules bind as a tetramer on one 9-nucleotide sequence.
DnaC – binds after DnaA is bonded and acts as a “matchmaker”, because by associating to OriC it allows DnaB to bind.
DnaB – DnaB is a helicase. Helicases are enzymes that separate the parental strands locally before the replication. Helicases bind to single stranded DNA (doesn't matter which one) and move along it in fixed direction, with each step requiring hydrolysis of ATP. This pushes the parental strand apart. The replication fork is created by DnaB.
ssDNA binding protein – after the two complementary strands of DNA have been separated by helicases, there is a big chance that they are going to form hydrogen bonds between each other, since they are complementary. A special protein called singe-stranded DNA binding protein, that has a big affinity for single stranded DNA, binds to the separated strands. It has the following functions:
prevents the DNA from reannealing
prevents the DNA from making hairpins
aligns the template strands for rapid DNA synthesis
falls off when DNA polymerase has passed over
These proteins are also important in recombination and repair.
Topoisomerase – Replication requires removal of positive supercoils created by overwinding of the helix immediately after the unwinding site.
In E.Coli:
Topoisomerase I removes negative supercoils, not sufficient for replication.
Topoisomerase II (gyrase) removes positive supercoils by inducing negative ones.
In human cells:
Topoisomerase I is capable of removing both negative and positive supercoils
Topoisomerase II is not a gyrase, but is essential for the termination step, segregation of chromosomes and appears to attach DNA to special sites to special site during interphase.
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