DNA Replication
The Specifics
- Before DNA replication can occur, highly coiled and condensed DNA molecule must be “straightened” into linear sequence of nucleotidesEnzymes serve this function.- Enzymes serve this function
DNA ReplicationThe Specifics
DNA Replication
DNA helicase
replication fork
Step 1: As DNA uncoils and unwinds, a class of enzymes called topoisomerases relieve tension on molecule
Step 2: DNA helicase breaks hydrogen bonds that hold two strands together - produces a “replication fork”
Topoisomerase
DNA Replication
Step 4: RNA primase attaches RNA nucleotides (primers) to 3´ end of each strand
RNA primers
3´
5´
3´5´
Single stranded binding proteins (SSBP) attach to each strand to prevent hydrogen bonds from reforming
Step 3:
SSBP
DNA ReplicationStep 5:
3´
5´
3´5´
DNA polymerase III5´
3´
5´
3´
Starting at primers DNA polymerase III adds complimentary DNA nucleotides to each strand
DNA polymerase III constructs new strand in 5
´-3´ direction only
DNA Replication3´
5´
3´5´
NOTICE:One strand (leading) is built continuously towards replication fork
Continuou
s
Other strand (lagging) is built discontinuously away from replication fork
Discontinuous
5´
3´5´
3´
Okazaki fragmentsLagging strand is made up
of short pieces called Okazaki fragments
DNA polymerase I replaces RNA primers with DNADNA ligase joins the Okazaki fragments.
Okazaki fragments
DNA Replication3´
5´
3´5´
Step 6:
DNA ligase
DNA ligase joins Okazaki fragments DNA polymerase I
Error Correction:Most occurs during Step 5 when DNA polymerase III is adding complimentary DNA nucleotides
Step 7:
T C
DNA Replication
T
AT CGAG GATG GCCCT
GACTC
3´5´5´3´ ACCGGGA
New strand is made from 5’ to 3’
1000 nucleotides added per second in bacteria
50 nucleotides added per second in Eukaryotes
When this happens, enzyme usually backs up and corrects error, before continuing on
Step 7: Error Correction:Most occurs during Step 5 when DNA polymerase III is adding complimentary DNA nucleotidesOccasionally, DNA polymerase III will make a mismatch error
DNA Replication
C T
AT CGAG GTG GCCCT
GACTC
3´5´5´3´ ACCGGGA
A
G
Uncorrected Mismatch Error
Step 7: Error Correction:Most occurs during Step 5 when DNA polymerase III is adding complimentary DNA nucleotidesOccasionally, DNA polymerase III will make a mismatch error
About one error in a million base pairs is not corrected by DNA polymerase III
When this happens, enzyme usually backs up and corrects error, before continuing on
DNA Replication
C CAC
T
AT CGAG GTG GCCCT
GACT
3´5´5´3´ CGGGA
G
A
Special DNA repair complexes made up of proteins and enzymes, including DNA polymerases I and DNA polymerase II, remove a small section of nucleotides around error
Step 7:
C
DNA Replication
CAC T
AT CGAG GTG GCCCT
GACT
3´5´5´3´ CGGGA
A
T
Special DNA repair complexes made up of proteins and enzymes, including DNA polymerases I and DNA polymerase II, remove a small section of nucleotides around error
Step 7:
Missing section is replaced by DNA polymerase IIId by DNA ligase.
C
DNA Replication
CAC T
AT CGAG GTG GCCCT
GACT
3´5´5´3´ CGGGA
A
T
This phosphodiester bond is missing
Missing phosphodiester bond of last nucleotide is joined by DNA ligase
Special DNA repair complexes made up of proteins and enzymes, including DNA polymerases I and DNA polymerase II, remove a small section of nucleotides around error
Step 7:
Missing section is replaced by DNA polymerase IIId by DNA ligase.
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