Active nucleoprotein filaments of single-stranded binding protein and recA protein on single-stranded DNA have a regular repeating structure.
When E. coli single-stranded DNA binding protein (SSB) coats single-stranded DNA (ssDNA) in the presence of 1 mM MgCl2 it inhibits the subsequent binding of recA protein, whereas SSB binding to ssDNA in 12 mM MgCl2 promotes the binding of recA protein. These two conditions correspond respectively to those which produce 'smooth' and 'beaded' forms of ssDNA-SSB filaments. By gel filtration and immunoprecipitation we observed active nucleoprotein filaments of recA protein and SSB on ssDNA that contained on average 1 monomer of recA protein per 4 nucleotides and 1 monomer of SSB per 20-22 nucleotides. Filaments in such a mixture, when digested with micrococcal nuclease produced a regular repeating pattern, approximately every 70-80 nucleotides, that differed from the pattern observed when only recA protein was bound to the ssDNA. We conclude that the beaded ssDNA-SSB nucleoprotein filament readily binds recA protein and forms an intermediate that is active in the formation of joint molecules and can retain substantially all of the SSB that was originally bound.
Muniyappa
K
K
Department of Human Genetics, Yale University School of Medicine, New Haven, CT 06510.
Williams
K
K
Chase
J W
JW
Radding
C M
CM
D002850:Chromatography, Gel; D004277:DNA, Single-Stranded / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000378:metabolism*; D007700:Kinetics; D015636:Magnesium Chloride / Q000494:pharmacology; D008836:Micrococcal Nuclease / Q000378:metabolism; D009698:Nucleoproteins / Q000378:metabolism*; D011233:Precipitin Tests; D011938:Rec A Recombinases / Q000378:metabolism*; D012091:Repetitive Sequences, Nucleic Acid
D004277:DNA, Single-Stranded; D004268:DNA-Binding Proteins; D009698:Nucleoproteins; D015636:Magnesium Chloride; D011938:Rec A Recombinases; D008836:Micrococcal Nuclease