DNA Structure and Topology
Understand the difference between the
major and minor groove. Specific DNA-binding proteins mainly contact DNA through hydrogen bonds in the major groove of B-DNA.
a) Why might sequence-specific binding be more common in the major groove than
in the minor groove?
b) What hydrogen-bond contacts can proteins make with the bases in the major
groove? Are these different from those in the minor groove?
Summarize the differences between A-,
B-, and Z-DNA. Under what solvent conditions would you expect
to find each one? What general sequence of bases is found in segments of DNA that are thought to be in the Z-DNA structure?
Be prepared to explain the various
forces involved in stabilizing DNA.
What are the mechanisms by which
flexibility of DNA is constrained?
Be able to recognize the various
parameters that contribute to DNA's
conformational flexibility if you are given structures. (ex:
c-angle, backbone torsion angles, syn-
and anti- steric orientations, nucleotide sugar
Thoroughly understand the mechanisms
of action of Type I (both IA and IB) and Type II Topoisomerases.
Distinguish between the
"controlled rotation" mechanism and the "strand passage"
mechanism and know which mechanism is used for Type IA, Type IB and Type II Topoisomerases.
Briefly explain the mechanisms of
action of Ciprofloxacin, Novobiocin, Etoposide, and Doxorubicin in blocking topoisomerase
Explain the following terms: twist,
writhe, super-coiling, linking number and be able to determine these for simple
models of DNA.
DNA negative supercoils can be titrated out by "unwinding agents." Ethidium bromide, a planar molecule,
"intercalates" between the stacked DNA bases, thereby unwinding the supercoils.
However, the linking number of the DNA is not changed. Explain the physical basis for the ability of ethidium bromide to "unwind" these supercoils.
Eukaryotic DNA does not have a DNA gyrase activity, as does
bacteria. How are negative supercoils
introduced into eukaryotic DNA such that the DNA can be compacted?
DNA Gyrase and DNA Topoisomerase IA have diametric functions in
E. coli. Explain.
You have discovered an enzyme
secreted by a particularly virulent bacterium that cleaves the C2'-C3' bond in
the deoxyribose residues of duplex DNA. What is the effect of this enzyme on supercoiled
DNA? Explain your answer.
A closed circular duplex DNA has a 100-bp segment of alternating C and G residues. On transfer to a
solution containing a high salt concentration, this segment undergoes a
transition from the B conformation to the Z conformation. What is the accompanying
change in its linking number, writhing number and twist?
Explain how negative supercoils can be introduced into eukaryotic DNA, despite the fact that eukarytic DNA do not have DNA Gyrase activity. I
may provide a diagram of the formation of one wrap of a circular DNA around a histone protein, followed by
removal of the compensatory (+) supercoil. You
will explain the steps involved, mentioning the enzyme type involved.
When the helix axis of a closed
circular duplex DNA of 2340 bp is constrained
to lie in the plane, the DNA has a twist of 212. When released, the DNA takes up its normal twist of 10.4 bp/turn. Indicate the values of "Lk", "Wr", and
"T" for both the constrained and unconstrained conformational states
of this DNA circle.