Human Rad51 mediated DNA unwinding is facilitated by conditions that favour Rad51-dsDNA aggregation.
BACKGROUND
Human Rad51 (RAD51), analogous to its bacterial homolog, RecA, binds and unwinds double stranded DNA (dsDNA) in the presence of certain nucleotide cofactors. ATP hydrolysis is not required for this process, because even ATP non hydrolysable analogs like AMP-PNP and ATPgammaS, support DNA unwinding. Even ADP, the product of ATP hydrolysis, feebly supports DNA unwinding.


RESULTS
We find that human Rad52 (RAD52) stimulates RAD51 mediated DNA unwinding in the presence of all Adenine nucleotide cofactors, (except in AMP and no nucleotide conditions that intrinsically fail to support unwinding reaction) while enhancing aggregation of RAD51-dsDNA complexes in parallel. Interestingly, salt at low concentration can substitute the role of RAD52, in facilitating aggregation of RAD51-dsDNA complexes, that concomitantly also leads to better unwinding.


CONCLUSIONS
RAD52 itself being a highly aggregated protein perhaps acts as scaffold to bring together RAD51 and DNA molecules into large co-aggregates of RAD52-RAD51-DNA complexes to promote RAD51 mediated DNA unwinding reaction, when appropriate nucleotide cofactors are available, presumably through macromolecular crowding effects. Our work highlights the functional link between aggregation of protein-DNA complexes and DNA unwinding in RAD51 system.
Balakrishnan
Kamakshi
K
Department of Biological Sciences, Tata Institute of Fundamental Research, Homi Bhabha Road, Colaba, Mumbai-400 005, India. kamakshihema@tifr.res.in
Krishnan
Neeraja M
NM
Kulkarni
Anagha
A
Rao
Basuthkar J
BJ
D004247:DNA / Q000378:metabolism*; D004260:DNA Repair; D004264:DNA Topoisomerases, Type I / Q000378:metabolism; D006801:Humans; D009711:Nucleotides / Q000494:pharmacology; D011189:Potassium Chloride / Q000737:chemistry / Q000378:metabolism; D051135:Rad51 Recombinase / Q000378:metabolism*; D051721:Rad52 DNA Repair and Recombination Protein / Q000378:metabolism*; D013997:Time Factors
D009711:Nucleotides; C497194:RAD52 protein, human; D051721:Rad52 DNA Repair and Recombination Protein; D011189:Potassium Chloride; D004247:DNA; C495153:RAD51 protein, human; D051135:Rad51 Recombinase; D004264:DNA Topoisomerases, Type I