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Quiz

BIOL 239 Quiz: Set 11- DNA Replication
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8 Pages
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Department
Biology
Course Code
BIOL239
Professor
Christine Dupont

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DNA Replication
The Watson-crick DNA replication model
oThis model proposed a mechanism
whereby DNA replication is
semiconservative: in daughter molecules one strand is conserved
from the parental molecule and the
other is newly synthesized
oDuring replication the strands
separate and used as templates to
make two new daughter strands
What are the three possible models of DNA replication?
oSemiconservative:
DNA molecules split and daughter molecules
form one new strand one old strand
oConservative:
DNA molecule makes two new molecules by itself
oDispersive:
Parts of a new molecule is dispersively used to
form old bits from new bits
Error! Filename not specified.
Who proved DNA semi-conservation? How?
oMatthew Meselson and Franklin Stahl
Experimental proof of semiconservative replication
oStudies with controlled isotopic composition
of nucleotides incorporated into daughter DNA strands
oGrew cells with N14 isotope and N15 isotope
so they are incorporated into their DNA.
oDNA was centrifuged so that heavier DNA (N15) was at
the core of tube and lighter DNA (N14) was closer
to the tip.
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o Hybrid DNA was in-between these two areas.
oBy looking at the banding
patterns could rule out the
conservative model as you
would see two clear N15 and
N14 bands
oTo rule out the semiconservative
model of DNA replication they let the
replicated strands to re-replicate
and found that they saw ¼ of the
progeny DNA hybrid and ¾ light
(N14)
The molecular mechanism of DNA
replication
oComplex process which occurs at
a precise moment is cell cycle
oTwo steps: initiation phase and
elongation (copying)
oE. coli DNA replication as a model
The reason E. coli is a very important model organism is because it is very
similar
Difference between prokaryote and eukaryote DNA?
oProkaryotes: Small circular (binary
fission)
Eukaryotes: Large linear
Describe the Initiation phase
oEukaryotes have this at s phase
oOrigin of replication = OriC
oOriC is at a site of A-T base pairs since hydrogen bonds on these bases are easier to
pull apart from each other
oInitiator proteins: Proteins that
recognize the sequence and the
origin and start the strand separation
of the two strands
oDNA Helicase: One one the first
enzymes acting on the replication
bubble and works to unwind the
DNA strand and works in both
directions of the replication bubble
oWhen this bubble gets to a certain
size as it gets bigger, DNA starts to
base pair together as this is the
energetically favorable reaction.
oSingle-stranded- DNA- binding proteins: Stabilize the single strands and stop them
from re-pairing together
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Elongation:
oDNA polymerase
(the enzyme that
builds DNA)
MUST have free
3’ OH to extend
upon and
requires a primer
to begin synthesis
oDNA primase: An enzyme that builds
RNA primer which provides a free 3’ OH
which can be extended on by DNA
polymerase
oAs helicase continues to make the
bubble bigger, DNA Polymerase
III starts to add nucleotides to the
3’OH end of the new strands in the
5’ to 3’ direction
oLeading strand: continuous
synthesis happens here as the
bubble opens up as DNA
polymerase III synthesizes DNA in
the 3’ to 5’ direction
oLagging strand: The discontinuous
synthesis of DNA occurs on the
lagging strand due to the direction
of the synthesis of DNA
polymerase III being in the 3’ to 5’
direction
oOkazaki fragments: DNA- RNA strands that make up the lagging strands,
which are small fragments of DNA due to the synthesis of the lagging strand
oDuring elongation, the RNA primers are removed by DNA polymerase I
oDNA polymerase 1 fills in the gap (still 5’ to 3’)
oDNA ligase seals the 3’ OH and 5’ PO4 nicks by catalyzing the formation of
phosphodiester bonds
oThis completes the formation of the lagging strand
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Description
DNA Replication  The Watson-crick DNA replication model o This model proposed a mechanism whereby DNA replication is semiconservative: in daughter molecules one strand is conserved from the parental molecule and the other is newly synthesized o During replication the strands separate and used as templates to make two new daughter strands  What are the three possible models of DNA replication? o Semiconservative:  DNA molecules split and daughter molecules form one new strand one old strand o Conservative:  DNA molecule makes two new molecules by itself o Dispersive:  Parts of a new molecule is dispersively used to form old bits from new bits Error! Filename not specified.  Who proved DNA semi-conservation? How? o Matthew Meselson and Franklin Stahl Experimental proof of semiconservative replication o Studies with controlled isotopic composition of nucleotides incorporated into daughter DNA strands o Grew cells with N14 isotope and N15 isotope so they are incorporated into their DNA. o DNA was centrifuged so that heavier DNA (N15) was at the core of tube and lighter DNA (N14) was closer to the tip. o Hybrid DNA was in-between these two areas. o By looking at the banding patterns could rule out the conservative model as you would see two clear N15 and N14 bands o To rule out the semiconservative model of DNA replication they let the replicated strands to re-replicate and found that they saw ¼ of the progeny DNA hybrid and ¾ light (N14)  The molecular mechanism of DNA replication o Complex process which occurs at a precise moment is cell cycle o Two steps: initiation phase and elongation (copying) o E. coli DNA replication as a model  The reason E. coli is a very important model organism is because it is very similar  Difference between prokaryote and eukaryote DNA? o Prokaryotes: Small circular (binary fission) Eukaryotes: Large linear  Describe the Initiation phase o Eukaryotes have this at s phase o Origin of replication = OriC o OriC is at a site of A-T base pairs since hydrogen bonds on these bases are easier to pull apart from each other o Initiator proteins: Proteins that recognize the sequence and the origin and start the strand separation of the two strands o DNA Helicase: One one the first enzymes acting on the replication bubble and works to unwind the DNA strand and works in both directions of the replication bubble o When this bubble gets to a certain size as it gets bigger, DNA starts to base pair together as this is the energetically favorable reaction. o Single-stranded- DNA- binding proteins: Stabilize the single strands and stop them from re-pairing together  Elongation: o DNA polymerase (the enzyme that builds DNA) MUST have free 3’ OH to extend upon and requires a primer to begin synthesis o DNA primase: An enzyme that builds RNA primer which provides a free 3’ OH which can be extended on by DNA polymerase o As helicase continues to make the bubble bigger, DNA Polymerase III starts to add nucleotides to the 3’OH end of the new strands in the 5’ to 3’ direction o Leading strand: continuous synthesis happens here as the bubble opens up as DNA polymerase III synthesizes DNA in the 3’ to 5’ direction o Lagging strand: The discontinuous synthesis of DNA occurs on the lagging strand due to the direction of the synthesis of DNA polymerase III being in the 3’ to 5’ direction o Okazaki fragments: DNA- RNA strands that make up the lagging strands, which are small fragments of DNA due to the synthesis of the lagging strand o During elongation, the RNA primers are removed by DNA polymerase I o DNA polymerase 1 fills in the gap (still 5’ to 3’) o DNA ligase seals the 3’ OH and 5’ PO ni4ks by catalyzing the formation of phosphodiester bonds o This completes the formation of the lagging strand o This illustration shows the synthesis and replacement of RNA primers during replication of the lagging strand of DNA. o A short RNA strand is synthesized to provide a 3’-OH primer for DNA synthesis. The RNA primer is subsequently removed and replaced with DNA by the dual 5’→ 3’ exonuclease and 5’ → 3’ polymerase activities built into DNA polymerase I. DNA ligase then covalently closes the nascent DNA chain, catalyzing the formation of phosphodiester linkages between adjacent 3’-hydroxyls and 5’-phosphates. o The 5’ → 3’ exonuclease activity of DNA polymerase I excises the RNA primer, and, at the same time, the 5’ → 3’ polymerase activity of the enzyme replaces the RNA with a DNA chain by usin
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