Biology Reference
In-Depth Information
30%.
113,114
The NPF exists in the collapsed configuration with ADP.
115
The extended conformation is believed to facilitate homology sampling on
duplex DNA. The ATP hydrolysis rate of RAD51 is severalfold slower com-
pared to the bacterial RecA.
116
However, this hydrolysis is still important for
the dissociation of recombinase from both newly formed heteroduplex DNA
and fortuitously bound RAD51 on duplex DNA.
85
The RAD51 NPF is not a static structure. Breast cancer susceptibility gene
product 2 (BRCA2) is known to nucleate RAD51 NPF formation on ssDNA at
the dsDNA-ssDNA junction.
117-120
Many helicases in yeast and humans
have been identified both
in vivo
and
in vitro
that act as anti-recombinases
capable of dissociating RAD51 from ssDNA. These include Srs2 in yeast, and
BLM and RECQ5 in humans.
68,121-124
Biophysical studies also indicate that
Srs2 augments the Rad51 ATPase activity within the NPF to facilitate rapid
protein turnover.
121
Moreover, both ensemble and single-molecule experiments
show a direct correlation of recombinase turnover from DNA and its ATPase
activity.
99,100,110,125-127
Homology search during the synapsis phase by RAD51/RecA family is by
random collision that involves transient nonspecific interactions with dsDNA,
presumably bound at the secondary DNA-binding site.
99,127
The transient
triplex DNA formed during the homology search is paramenic and not topo-
logically interwound.
99
Biophysical studies using fluorescence resonance ener-
gy transfer and selective substitution of guanine to inosine on both ssDNA and
its identical strand on the duplex DNA showed that human RAD51 facilitated
homology search by a rapid A:T base-flipping mechanism.
128
Once a homolo-
gous sequence is found, strand exchange occurs to produce a topologically
interwound plectonemic heteroduplex DNA product.
99,127
by
B. DMC1
Disrupted meiotic cDNA (DMC1) was first isolated from a budding yeast
meiotic cDNA library screen.
129
DMC1 is only expressed during meiosis and
displays considerable homology to the RecA/RAD51 family of recombinases
(
Table I
; Ref.
129
). During meiosis, both Rad51 and Dmc1 colocalize at DSBs
in yeast.
130
Disruption of yeast Dmc1 leads to a number of abnormal meiotic
phenotypes, including accumulation of DSBs, reduced reciprocal recombina-
tion, abnormal synaptonemal complex formation, and defective meiotic pro-
phase arrest.
129
Dmc1 exhibits both overlapping and yet nonredundant
functions to Rad51.
131
However, overexpression of Rad51 enables yeast cells
to circumvent the defective meiotic phenotype of Dmc1 mutants.
131
Human
DMC1 exists as an octamer in solution
132
and functions similar to RAD51 in
assays for recombination and ATPase activity
in vitro
.
126,133,134
