DNA replication in Prokaryotes complete notes
DNA replication in prokaryotes
• The circular chromosomal DNA of E.coli is replicated bidirectionally from a single origin of replication, oriC.
• This mode of replication is also known as θ-replication.
• This process occurs in three stages.
- Initiation
- Elongation
- Termination
Initiation
This involves recognition of the position on a DNA
molecule where replication will begin.
Elongation
This involves the events occuring at the replication fork,
where the parent DNA strands are copied.
Termination
This occurs when the parent DNA molecule has been completely
replicated.
Initiation of DNA replication in E.coli
·
DnaA protein initiates replication by binding to the oriC.
·
Binding of DnaA facilities the melting of duplex DNA.
·
Further melting is mediated by DnaB protein ( Helicase
) which is accompanied by DnaC protein ( Helicase loader )
· The separated strands are inhibited from reannealing by Single-stranded binding protein ( SSB ). It binds to both strands separately.
·
DnaB protein ( Helicase ) recruits a DnaG enzyme ( DNA
primase ) that synthesize an RNA primer ( short RNA molecule ) on
each template.
· The RNA primer causes
the DnaC protein ( DNA helicase loader ) to release from DnaB
protein ( helicase ).
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Elongation
·
DNA polymerase 3 catalyzes the step by step addition of deoxyribonucleotide
units to a growing DNA chain.
·
DNA replication of E.coli chromosomal DNA is bi-directional
& semi-discontinuous.
·
In the case of bi-directional replication , two
replication forks form that move in opposite directions.
· In the case of semi-discontinuous replication , one
strand ( the leading strand ) replicates continuously & the
other strand ( the lagging strand ) replicated discontinuously as
okazaki fragments.
·
As each okazaki fragment formation completes , the RNA primer
of the fragment is removed by DNA polymerase 1.
·
Removal of the RNA primer leaves a gap in the dsDNA.
·
DNA polymerase 1 also fills the gaps b/w the lagging strand fragments.
·
After the replacement of all the RNA primers with DNA , all of
the separately primed lagging-strand DNA fragments are joined
together to form one continuous DNA strand by enzyme DNA ligase.
As the strands of DNA are
separated at the replication fork due to DNA helicase , the dsDNA in front of
the fork becomes increasing positively supercoiled. In E.coli DNA
gyrase is required to relieve this positive supercoiling
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Termination
·
Replication of genome
terminates at terminus region containing multiple copies of ter (
terminus sequences ).
·
Several ter sequences acts as the recognition site for a Tus
( terminus utilization substance ) protein.
·
The orientation of the ter sequences & hence
of the bound Tus proteins is such that both replication forks becomes
trapped on the opposite side of the origin of replication.
·
When replication is finished , the two new circular
chromosome may be physically interlocked or catenated. Such
interlocked DNA must be separated by DNA topoisomerase 4.
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