Purification of a RecA protein analogue from Bacillus subtilis.
We have identified in Bacillus subtilis an analogue of the Escherichia coli RecA protein. Its activities suggest that it has a corresponding role in general genetic recombination and in regulation of SOS (DNA repair) functions. The B. subtilis protein (B. subtilis Rec) has a Mr of 42,000 and cross-reacts with antisera raised against E. coli RecA protein. Its level is significantly reduced in the recombination-deficient recE4 mutant. B. subtilis Rec is induced 10- to 20-fold in rec+ strains following treatment with mitomycin C, whereas it is not induced in the recombination-deficient mutants recE4, recE45, and recA1. We have purified B. subtilis Rec about 2000-fold to near homogeneity and we describe its activities. It catalyzes DNA-dependent hydrolysis of dATP at a rate comparable to that of E. coli RecA protein. However, B. subtilis Rec has a negligible ATPase activity, although ATP effectively inhibits dATP hydrolysis. In the presence of dATP, B. subtilis Rec catalyzes DNA strand transfer, assayed by the conversion of phi X174 linear duplex DNA and homologous circular single-stranded DNA to replicative form II (circular double-stranded DNA with a discontinuity in one strand). ATP does not support strand transfer by this protein. B. subtilis Rec catalyzes proteolytic cleavage of E. coli LexA repressor in a reaction that requires single-stranded DNA and nucleoside triphosphate. This result suggests that an SOS regulatory system like the E. coli system is present in B. subtilis. The B. subtilis enzyme does not promote any detectable cleavage of the E. coli bacteriophage lambda repressor.
Lovett
C M
CM
Roberts
J W
JW
D000251:Adenosine Triphosphatases / Q000378:metabolism; D001412:Bacillus subtilis / Q000032:analysis*; D001426:Bacterial Proteins / Q000378:metabolism; D002499:Centrifugation, Density Gradient; D003429:Cross Reactions; D004277:DNA, Single-Stranded / Q000378:metabolism; D003838:Deoxyadenine Nucleotides / Q000378:metabolism; D004587:Electrophoresis, Agar Gel; D004591:Electrophoresis, Polyacrylamide Gel; D016685:Mitomycin; D008937:Mitomycins / Q000494:pharmacology; D011938:Rec A Recombinases / Q000032:analysis*; D012697:Serine Endopeptidases; D000099160:LexA Repressor Protein
D000251:Adenosine Triphosphatases; D001426:Bacterial Proteins; D004277:DNA, Single-Stranded; D003838:Deoxyadenine Nucleotides; D016685:Mitomycin; D008937:Mitomycins; D011938:Rec A Recombinases; D012697:Serine Endopeptidases; D000099160:LexA Repressor Protein; C026600:2'-deoxyadenosine triphosphate