DNA REPLICATION Flashcards
What is semiconservative replication of DNA?
the 2 original strands of DNA separate and each acts as a template on which a new complementary strand is laid down
(storage and transmission of genetic information)
Who proved DNA replication is semiconservative?
Meselson and Stahl (1957)
used ecoli and heavy nitrogen
What did Kornberg do in 1958?
descried the purification of an enzyme called DNA polymerase I from Ecoli that synthesized DNA
How is DNA synthesized?
from nucleotide triphosphate precursors by the enzyme DNA polymerase
What did John Cairns in 1963 reveal?
that an ecoli chromosome is a single 1.7mm circular molecule (replicating DNA looks like theta)
How is DNA replicated in bacteria? (remember circular structure)
bidirectionally because a replication fork is found on each end of the loop where DNA is unwound and the daughter strand is being synthesized
(don’t worry too much about this)
What is a primer?
a segment of nucleic acid (RNA or DNA) base paired to the template and with a free 3’-OH
Who discovered the DNA had a template strand and base pairing rules?
Watson-Crick
What is processivity?
number of bases synthesized before the DNA polymerase enzyme dissociates from the template (can be thousands)
What does DNA polymerase exhibit?
processivity
Is DNA polymerase and DNA replication accurate?
yes very accurate
Is DNA replication more accurate in humans or bacteria?
bacteria
What improves accuracy of DNA polymerase and DNA replication?
mismatch repair
What causes tautomers to form?
chemical structure is not absolutely stable, proton rearrangement leads to different chemical forms
What are tautomers?
the same atomic composition with a slightly different chemical structure (changes the pairing characteristics of specific bases)
At any given time roughly how many tautomers are there?
1 in 10,000 molecules are in rare tautomeric form
A cytosine tautomer base pairs accurately with what?
adenine (instead of guanine like it would if it was not a tautomer)
What is the proofreading activity of DNA polymerase I?
3’ to 5’ exonuclease activity
acts ONLY on the base just incorporated (no others)
Does polymerization activity stop when there is an error found?
yes because it has to stop and fix the problem (remove mispaired nucleotide) then it will resume
How many E coli DNA polymerases are there and what are they?
3
DNA polymerase I
DNA polymerase II
DNA polymerase III
What is the function of DNA polymerase I in Ecoli?
mostly for repair
What is the function of DNA polymerase II in Ecoli?
(specialized repair function) don’t need to know
What is the function of DNA polymerase III in Ecoli?
mismatch repair (main DNA replication enzyme)
Which DNA polymerase in Ecoli is very processive?
DNA polymerase III
once it clamps on the DNA it stays on for a long time and adds over half a million bases before it is removed
How many subunits does DNA polymerase III have?
10
What are the 3 genes that encode the 3 DNA polymerases?
polA (correlates with DNA polymerase I), polB (correlates with DNA polymerase II), polC(correlates with DNA polymerase III)
What is the activity of polA and what exactly does it do?
has a 5’ to 3’ exonuclease activity used to get rid of primer RNA during DNA replication
Is polA, polB, or polC the large gene that makes a big donut shaped clamp?
polC
What are the 3 stages of DNA replication?
initiation
elongation
termination
Which stage in DNA replication most complicated?
initiation
Where does DNA replication begin?
origin of replication (ORI)
How many proteins are involved in DNA replication in Ecoli and is it more or less than in humans?
about 20 which is more than what is used for humans
What is DUE? (in Ecoli)
a 13 base pair sequence that unwinds the DNA to start the process of replication
What does it mean when you see GATC during DNA replication in bacteria?
means mismatch repair is taking place at the methylation site which tells us that replication has been initiated at this site (THIS WILL ONLY HAPPEN ONCE)
What are R region (R1-R5) seen in DNA replication in Ecoli?
a DnaA protein binding site that has five regions with a 9 base pair sequence
What happens at the R regions (R1-R5)?
DnaA protein binds at these 5 sites and causes DUE to open up into single strand portions