Prokaryotic DNA Replication
? Prokaryotic DNA Replication
DNA replication is perfomed by a multienzyme complex >1 MDa
DNA Nucleotides Replisome: DNA polymerases Helicase Primase SSBs DNA ligase Clamps (Topoisomerases)
1
Replication is semiconservative, accurrate and fast
Accuracy 1 error in 1 billion bases Speed 500 nt/s in bacteria 50 nt/s in mammals
Each original strand functions as template for DNA synthesis
2
After each replication cycle, DNA is doubled
DNA is synthesized in 5?to 3?direction
3
Polymerisation in detail
(dNMP)n + dNTP DNA
(dNTP)n+1 + PPi 2 Pi
Complementary basepairing and matching hydrogen bonds is required
Incorrect basepairing
4
DNA is synthesized by DNA polymerase
DNA polymerase III is a protein complex
Subunit
function
not known 3' exonuclease polymerase clamp dimerisation
clamp loader
5
E. coli contains multiple DNA polymerases
DNA pol I
DNA pol II
DNA pol III
Number/cell
400
100
10
Speed (nt/s)
16-20
2-5
250-1000
3?exonuclease Yes
Yes
No
5?exonuclease Yes
No
No
Processivity
3-200
10 000
500 000
Role
DNA repair
DNA repair
Replication
RNA primer
removal
DNA polymerase I
Found by Arthur Kornberg, mid 1950's
Three enzymatic activities:
?
Polymerase activity
?
3' to 5' exonuclease activity
?
5' to 3' exonuclease activity
Klenow enzyme is lacking one subunit responsible for the 5' to 3' exonuclease activity
6
DNA polymerase requires
1. A free 3'-OH group supplied by RNA Primer for start of polymerisation
2. Mg2+ ions for activity in active site 3. A template to copy
DNA replication initate at origin of replication
Bacterial chromosome doubles in 40 min
7
DNA replication is bidirectional
The replication origin OriC in E.coli
245 base pairs AT-rich Initiation proteins bind to 9 bp consensus sequence
8
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