Process of DNA Replication

1st step: Initiation

 The first stage in replication is to separate the two DNA strands of the parental DNA molecule.

 DNA replication begins at a origin of replication (oriC) and proceeds bidirectionally.

 During initiation, proteins bind to the origin of replication

 Helicase unwinds the DNA helix

 Two replication forks are formed at the origin of replication.

 During replication, the DNA strands of the double helix must be both unwound and separated.

 To unwind the DNA, positive superhelical turns have to be removed by cutting the DNA, and
allowing it to relax or by introducing negative superhelical turns to compensate for the positive
ones.
 Replication fork:

Region where the enzymes replicating

a DNA molecule are bound to untwisted,
single-stranded DNA
 2nd step: Elongation

 During elongation, an enzyme called DNA polymerase adds DNA nucleotides to the 3′ end of the
newly synthesized polynucleotide strand.

 DNA polymerase will only synthesize DNA in a 5’ to 3’ direction.

 DNA polymerases lack the ability to initiate a new strand and can only elongate a pre-existing
strand.

 Unlike DNA polymerases, RNA polymerases can start new strands without any pre-existing 3’-
OH. A special RNA polymerase, known as primase, makes the RNA primers that are responsible
for strand initiation during DNA synthesis.
Leading strand :

The “leading strand” is synthesized continuously toward the replication fork as helicase unwinds the
template double-stranded DNA. The leading strand only needs to be started once.

Lagging strand:

The “lagging strand” is synthesized in the direction away from the replication fork and away from the
DNA helicase unwinds. the lagging strand is made in short sections and a new RNA primer must be
created each time a new portion is made. DNA polymerase then builds new strands of DNA starting
from each RNA primer
Okazaki fragments

 Okazaki fragments are short sequences of DNA nucleotides which are synthesized
discontinuously and later linked together by the enzyme DNA ligase.

 These are1000-2000 bases in prokaryotes

 100-200 bases in eukaryotes.

 3rd step: Termination

 Termination of DNA replication occurs when two oppositely orientated replication forks
meet and fuse, to create two separate and complete double ‐stranded DNA molecules.

 Failure to terminate bacterial chromosome replication correctly results in chromosome

over‐replication and genome instability.
Which protein is used for termination of replication?

 The replication termination protein (RTP) is one of only two well-defined proteins known
to be involved in arresting DNA replication forks.

 the other being a protein known as tus (termination utilisation substance) from E. coli.