Central Dogma of Molecular

Biology
DNA Replication and
General Chracteristics
DNA Replication and
Proofreading in Prokaryotes
 The central dogma of molecular biology
describes the flow of genetic information in
cells from DNA to messenger RNA (mRNA) to
protein.
 It occurs in all Living Organisms.
 It is non-specific. It means that once DNA
replication begins, it will not stop (normally)
until all the DNA is replicated.
 It is semi-conservative.
 It is bidirectional.
 DNA polymerases read the template strand
from 3’ 5’ and synthesize the new
DNA strand from 5’ 3’.
 Initiation
DNA A box and DNA A Proteins
Formation of Replication Bubble and Replication Fork
SSB
Helicases
 Supercoiling and Topoisomerases
 Elongation
Elongation of Leading Strand
ProofReading
Okazaki fragments and Lagging Strand
DNA pol I and Ligases
 Termination
 DNA replication begins at a single,unique AT-
rich sequence site called “Origin of
Replication” or “Ori”.
 As the two strands unwind and separate ,
synthesis occurs bidirectionally generating a
replication bubble.
 Initiation of DNA replication requires a group
of proteins collectively called “Pre-Priming
Complex”.
 DnaA proteins binds to DnaA Boxes within origin
of replication , causing the short AT-rich regions
in the origin to melt.Melting is ATP dependent ,
and results in strand separation with the
formation of localized regions of ssDNA.
 Helicases bind to ssDNA near the replication fork
and then move into the neighboring double-
stranded region, forcing the strands apart.Energy
is provided by ATP.
 Unwinding causes the problem of supercoiling in
other regions of DNA molecule that is solved by
Topoisomerases.
 SSB proteins bind to
ssDNA generated
by helicases. They
keep the two
strands of DNA
separated and also
protect DNA from
Nucleases.
 Binding of SSB
proteins is Co-
operative.
 The strand that is being copied in the
direction of the advancing replication fork is
called the Leading strand and is synthesized
continuously.
 The strand that is being copied in the
direction away from replication fork is
synthesized discontinously ( okazaki
fragments) is called the Lagging Strand.
 Primase synthesizes the short stretches of
RNA that are complementary and antiparallel
to the DNA template. These are called Primer
sequence.
 Only one Primer sequence is required at the
Leading strand whereas primer sequences are
synthesized constantly at Lagging strand.
 After the synthesis of Primer , DNA Pol III
begins to add nucleotides along the single
stranded template.
 At the leading strand, nucleotides are being
added continously.
 At the lagging strand, DNA pol III adds the
nucleotides until it reaches the next primer
sequence. It then deattaches , and bind again
at the next primer sequence , causing it’s
elongation and formation of Okazaki
fragments.
 DNA pol III while synthesizing the new DNA
strands also Proofreads them by it’s 3’ 5’
exonuclease activity.
 It is highly important for the survival of an
organism that the nucleotide sequence of
DNA be replicated with as few errors as
possible.
 DNA pol I with it’s 5’ 3’ exonuclease
activity removes the primer sequences and
replaces them with DNA .
 The final phosphodiester linkage between the
DNA chain synthesized by DNA pol III and the
chain made by DNA pol I is joined by DNA
ligases.
 Termination in E.Coli is mediated by binding
of the protein “Tus (terminal utilization
substance)” to termination sites on the DNA ,
stopping the movement of DNA polymerase.