Chemistry Reference
In-Depth Information
were made from
five different DNA sequences implying that the result is not speci
c
to a particular sequence. The bias between the two adjacent branch points can be as
high as 40-fold, a surprising result considering the number of base pairs and their
nature (GC versus ATetc.) do not change after branch migration. This suggests that
local branch migration may be much faster than previously thought, and the
ensemble estimate of the branch migration rate must be affected greatly by the
presence of ultra-stable branch points. This also raises the possibility that spontane-
ous branchmigration of a transiently protein-free HJ in vivo can be substantial so that
these stable branch points are preferentially accessed by junction-resolving enzymes
or other recombination machinery.
11.5
Outlook
I have summarized some of the key
findings from our single-molecule FRETstudies
of helicase andHolliday junction. We were very lucky in that previous biophysical and
biochemical studies, for example by our collaborators Tim Lohman and David Lilley,
guided the design and interpretation of our single-molecule experiments to place our
results on
firm ground. Despite the extensive knowledge base previously available,
our single-molecule data provided some remarkable surprises, for example that a
helicase protein can repetitively shuttle on the same DNA segment. One immediate
extension would be to study the interaction between the helicases and Holliday
junctions. In fact, many recombination proteins that catalyze branchmigration of HJ
are classi
ed as helicases and it will be very interesting to investigate how the unique
properties of the HJ revealed by the single-molecule studies can in
uence the
activities of enzyme that recognize and act on the HJ.
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