Srs2 prevents Rad51 filament formation by repetitive motion on DNA.
Srs2 dismantles presynaptic Rad51 filaments and prevents its re-formation as an anti-recombinase. However, the molecular mechanism by which Srs2 accomplishes these tasks remains unclear. Here we report a single-molecule fluorescence study of the dynamics of Rad51 filament formation and its disruption by Srs2. Rad51 forms filaments on single-stranded DNA by sequential binding of primarily monomers and dimers in a 5'-3' direction. One Rad51 molecule binds to three nucleotides, and six monomers are required to achieve a stable nucleation cluster. Srs2 exhibits ATP-dependent repetitive motion on single-stranded DNA and this activity prevents re-formation of the Rad51 filament. The same activity of Srs2 cannot prevent RecA filament formation, indicating its specificity for Rad51. Srs2's DNA-unwinding activity is greatly suppressed when Rad51 filaments form on duplex DNA. Taken together, our results reveal an exquisite and highly specific mechanism by which Srs2 regulates the Rad51 filament formation.
Department of Bioengineering, University of Illinois at Urbana-Champaign, Urbana, Illinois 61801, USA.
Yupeng
Y
Qiu
Edwin
E
Antony
Sultan
S
Doganay
Hye Ran
HR
Koh
Timothy M
TM
Lohman
Sua
S
Myong
D000255:Adenosine Triphosphate / Q000378:metabolism; D004265:DNA Helicases / Q000235:genetics / Q000378:metabolism*; D004271:DNA, Fungal / Q000235:genetics / Q000378:metabolism*; D004277:DNA, Single-Stranded / Q000235:genetics / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D059765:Homologous Recombination / Q000235:genetics*; D011485:Protein Binding / Q000235:genetics; D051135:Rad51 Recombinase / Q000096:biosynthesis / Q000235:genetics*; D011938:Rec A Recombinases / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins / Q000096:biosynthesis / Q000235:genetics* / Q000378:metabolism*
D004271:DNA, Fungal; D004277:DNA, Single-Stranded; D004268:DNA-Binding Proteins; D029701:Saccharomyces cerevisiae Proteins; C061376:SRS2 protein, S cerevisiae; D000255:Adenosine Triphosphate; C495151:RAD51 protein, S cerevisiae; D051135:Rad51 Recombinase; D011938:Rec A Recombinases; D004265:DNA Helicases