Nucleic Acids Research, 1990, Vol. 18, No. 13 3967-3973
© 1990
MOLECULAR BIOLOGY |
Active nucleoprotein filaments of single-stranded binding protein and recA protein on single-stranded DNA have a regular repeating structure
1Department of Human Genetics, Yale University School of Medicine New Haven, CT 06510 2Molecular Biophysics and Biochemistry, Yale University School of Medicine New Haven, CT 06510 3Department of Molecular Biology, Albert Einstein College of Medicine Bronx, NY 10461 USA
*To whom correspondence should be addressed at Department of Biochemistry, Indian Institute of Science, Bangalore-560012, India
Received January 25, 1990. Revised May 5, 1990. Accepted May 5, 1990.
When E. coli single-stranded DNA binding protein (SSB) coats single-stranded DNA (ssDNA) in the presence of 1 mM MgCI2 it inhibits the subsequent binding of recA protein, whereas SSB binding to ssDNA in 12 mM MgCI2 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.
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