Chapter 20
1) Understand semi-conservative mode of DNA replication. Know the difference
between conservative, semi-conservative and random modes of DNA replication.
Understand the Messelson and Stahl experiments. If DNA replicated by a
conservative or random mechanism, what would the Messelson and Stahl
experimental results look like?
2) Know that DNA replication is bi-directional and semi-discontinuous. Know what
this means.
4) Be able to draw one replication fork (no enzymes required, also don’t need to
draw the structure of individual nucleotides, just distinguish DNA strands as
lines). Be able to show the leading strand, the lagging strand, and Okazaki
fragments. Show the direction of replication of each strand. Distinguish the 5’ and
3’ ends of all DNA molecules. Show the direction of movement of the replication
fork.
5) Know the functions of the three DNA polymerases in E. coli
a. DNA polymerase I – DNA repair and participates in synthesis of lagging
strand
b. DNA polymerase II – DNA repair
c. DNA polymerase III – major polymerase involved in DNA replication.
6) Know the function of the following DNA polymerase III subunits
a. core complex
i)
3’ – 5- exonulcease
ii)
5’-3’ DNA polymerase
b. beta subunit – sliding DNA clamp
7) Know what occurs during the different phases of transcription
a. Initiation
i)
identification of origin of replication – a protein identifies
specific DNA sequence
ii)
A DNA helicase unwinds the double helix
iii)
Primasome complex binds to unwound region and
synthesizes RNA primer
iv)
Two DNA polymerase bind to the individual DNA strands at
the replication fork.
b. Elongation
i)
Elongation involves DnaB helicase unwinding, SSB binding
to keep strands separated.
ii)
Primasome complex continues to synthesize RNA primers
for lagging strand synthesis
iii)
DNA polymerase grinding away on both strands
iv)
topoisomerase relieves supercoiling at both replication forks.
v)
Know how the DNA polymerase releases and rebinds during
the synthesis of the lagging strand
vi)
Know how DNA polymerase and DNA ligase act to connect
Okazaki fragment during the replication of the lagging
strand.
I.
IX.
III.
IV.
DNA polymerase I has 5’-3’ enxonuclease
activity that degrades the RNA primer.
Then the DNA polymerase I has 5’3’polymerase activity resynthesizes a DNA
strand to replace the RNA primer.
DNA ligase connects adjacent fragments
DNA ligase used NAD+ in the ligase reaction in
a phosphoryl transfer type reaction (no redox
function)
c. Termination
i)
occurs at ter region (specific DNA sequence)
ii)
termination utilization substance (Tus) protein binds to ter region and
prevents replication fork from passing (inhibits helicase)
iii)
Know the functions of the different eukaryotic DNA polymerases
I.
Alpha – Primer synthesis and DNA repair
II.
Beta – DNA repair
III.
Gamma – Mitochondrial DNA replication
IV.
Delta – Leading and lagging strand synthesis,
and DNA repair
V.
Epsilon – Repair and gap filling on lagging
strand.
iv)
know that the proliferating cell nuclear antigen (PCNA) plays an
analogous role to the beta-subunit of the bacterial DNA polymerase.
8) DNA repair
a. excision repair
i.
know the enzymes involved.
ii.
Know how DNA damage by the deamination of cytosine to
form uracil is repaired
b. repair of thymine dimers
i)
know what thymine dimers are and how they form
ii)
know what enzyme (DNA photolyase) repairs damage and
how light is involved.