DNA REPLICATION
OriC. = - 5 binding sites (R1,R2,R3,R4,R5 (sites rich in A & T)) for key initiator proteins (DnaA)
- Region rich in A=T (DNA unwinding element, DUE)
- 3 additional Dna-binding sites (I1,I2,I3)(only when DnaA in complexed with ATP)
- Binding sites for FIS & IHF proteins
IMPORTANT: DnaA is crucial for initiation (AAA+ ATPase); it’s active form is bound to ATP,
the inactive one to ADP (ATP is slowly hydrolysed to ADP).
INITIATION:
8 DnaA proteins assemble on R & I sites on OriC. → Dna wraps around complex (right-handed
helix) → this favours opening at DUE region (IHF, HU, FIS stabilize loop)
DnaC (AAA+ ATPase) load DnaB HELICASE (hexamer) at DUE → DnaC-DnaB complex
activates DnaB HELICASE → DnaC hydrolyses ATP and detaches leaving DnaB to open the
Replication Fork and migrate along in the 5’→3’ direction.
Dna polymerase III binds to DnaB + SSB proteins bind to strands to stabilize + Dna GYRASE
relieves topological pressure.
IMPORTANT: initiation is the only phase known to be regulated
Dna pol III binding to Dna signals completion of initiation
Hda (AAA+ ATPase) protein binds to Dna pol III & interacts with DnaA → ATP hyd. To ADP
DnaA complex disassembles at origin
DAM METHYLASE methylates N6 position of adenine (palindromic sequence: (5’)GATC) → after
replication Dna is hemi-methylated and it sequesters to the plasma membrane and binds to SeqA
proteins → entire Dna needs to be methylated before it can begin another round of replication
ELONGATION:
PRIMASE (DnaG) synthesises PRIMER (Rna) on leading strand (primosome = DnaB (helicase) +
DnaG (primase))
, Leading strand: Dna pol III adds deoxyribonucleotides (Dna pol III linked to DnaB helicase on
opposite strand → leading strand synthesis proceeds continuously)
Lagging strand: primase synthesises Rna primer and Dna pol III adds deoxyribonucleotides → for
synthesis to occur on both strands the lagging strand must loop so that the 3 subunits of Dna pol III
can function (1 subunit on leading and 2 cycle continuously on lagging)
DnaB helicase moves along and unwraps replication fork; DnaG primase occasionally associated
and forma Rna primers → Dna pol III positions new beta sliding clamp → clamp-loading complex
is and AAA+ ATPase
Once an Okazaki fragment has been completed it’s primer is removed by Dna pol III and replaced
with Dna → nick is sealed by Dna LIGASE
TERMINATION:
TER sequence signals end of replication → binding sites for TUS protein → replication fork
arrested in 1 direction, other fork halts when it meets the first → TOPOISOMERASE IV separates
the 2 circular chromosomes (CATENATES, in E. Coli)
TELOMERES
Found at the end of linear Dna chromosomes: telomeres (double strands of deoxyribonucleic-
monophosphate-nucleotides) characterised by a series of repeated sequences.
TELOMERASE: enzyme (reverse transcriptase: Rna-dependent Dna polymerase), contains
oligoribonucleotide template (Rna template)
Elongates the template strand (leading strand, lagging strand leaves a gap at it’s the end after all of
the Rna primers have been removed) by binding to a recognition sequence and using its own
template to continue elongation.
Inverse correlation between telomere length and age (in somatic cells)
OriC. = - 5 binding sites (R1,R2,R3,R4,R5 (sites rich in A & T)) for key initiator proteins (DnaA)
- Region rich in A=T (DNA unwinding element, DUE)
- 3 additional Dna-binding sites (I1,I2,I3)(only when DnaA in complexed with ATP)
- Binding sites for FIS & IHF proteins
IMPORTANT: DnaA is crucial for initiation (AAA+ ATPase); it’s active form is bound to ATP,
the inactive one to ADP (ATP is slowly hydrolysed to ADP).
INITIATION:
8 DnaA proteins assemble on R & I sites on OriC. → Dna wraps around complex (right-handed
helix) → this favours opening at DUE region (IHF, HU, FIS stabilize loop)
DnaC (AAA+ ATPase) load DnaB HELICASE (hexamer) at DUE → DnaC-DnaB complex
activates DnaB HELICASE → DnaC hydrolyses ATP and detaches leaving DnaB to open the
Replication Fork and migrate along in the 5’→3’ direction.
Dna polymerase III binds to DnaB + SSB proteins bind to strands to stabilize + Dna GYRASE
relieves topological pressure.
IMPORTANT: initiation is the only phase known to be regulated
Dna pol III binding to Dna signals completion of initiation
Hda (AAA+ ATPase) protein binds to Dna pol III & interacts with DnaA → ATP hyd. To ADP
DnaA complex disassembles at origin
DAM METHYLASE methylates N6 position of adenine (palindromic sequence: (5’)GATC) → after
replication Dna is hemi-methylated and it sequesters to the plasma membrane and binds to SeqA
proteins → entire Dna needs to be methylated before it can begin another round of replication
ELONGATION:
PRIMASE (DnaG) synthesises PRIMER (Rna) on leading strand (primosome = DnaB (helicase) +
DnaG (primase))
, Leading strand: Dna pol III adds deoxyribonucleotides (Dna pol III linked to DnaB helicase on
opposite strand → leading strand synthesis proceeds continuously)
Lagging strand: primase synthesises Rna primer and Dna pol III adds deoxyribonucleotides → for
synthesis to occur on both strands the lagging strand must loop so that the 3 subunits of Dna pol III
can function (1 subunit on leading and 2 cycle continuously on lagging)
DnaB helicase moves along and unwraps replication fork; DnaG primase occasionally associated
and forma Rna primers → Dna pol III positions new beta sliding clamp → clamp-loading complex
is and AAA+ ATPase
Once an Okazaki fragment has been completed it’s primer is removed by Dna pol III and replaced
with Dna → nick is sealed by Dna LIGASE
TERMINATION:
TER sequence signals end of replication → binding sites for TUS protein → replication fork
arrested in 1 direction, other fork halts when it meets the first → TOPOISOMERASE IV separates
the 2 circular chromosomes (CATENATES, in E. Coli)
TELOMERES
Found at the end of linear Dna chromosomes: telomeres (double strands of deoxyribonucleic-
monophosphate-nucleotides) characterised by a series of repeated sequences.
TELOMERASE: enzyme (reverse transcriptase: Rna-dependent Dna polymerase), contains
oligoribonucleotide template (Rna template)
Elongates the template strand (leading strand, lagging strand leaves a gap at it’s the end after all of
the Rna primers have been removed) by binding to a recognition sequence and using its own
template to continue elongation.
Inverse correlation between telomere length and age (in somatic cells)
