RecA protein--promoted lambda repressor cleavage: complementation between RecA441 and RecA430 proteins in vitro.
Induction of prophage lambda occurs in recA441 mutant lysogens after a shift to 42 degrees C in the presence of adenine. If the synthesis of RecA441 protein is maintained at a low basal level by the presence of a second mutation in the recA441 gene, recA453, induction of prophage lambda is prevented. The ability to induce prophage lambda is restored by the introduction, on a transducing phage, of a second recA gene carrying the recA430 mutation; by itself, the RecA430 protein is devoid of activity against the lambda repressor (Rebollo et al. 1984). In order to explain how the RecA430 protein might complement the RecA441 protein to provide lambda repressor cleavage in a recA453-441 (recA430) diploid lysogen, we characterized the cleavage reaction catalysed by a mixture of these proteins in vitro. Our results suggest that, in the presence of dATP, the RecA441 and RecA430 proteins form mixed multimers on single-stranded DNA, in which the RecA441 protein molecules enhance the DNA binding affinity of RecA430 protein molecules, but RecA430 protein molecules support no cleavage of the lambda repressor. Although the effects of the RecA430 and single-strand binding (SSB) proteins are similar in vitro, we show that the SSB protein cannot substitute for the RecA430 protein in restoring lambda repressor cleavage in a recA453-441 lysogen. Comparison of the stimulatory effect of long single-stranded DNA with that of (dA)14 oligonucleotides on the RecA441 protein-directed cleavage of the lambda repressor in the presence of various nucleoside triphosphates (NTPs) indicates that the cooperative binding of the RecA441 protein to single-stranded DNA stabilizes the RecA protein-DNA complexes so that they remain intact long enough to support cleavage of the lambda repressor. We conclude that the low basal level of the RecA441 protein in a recA453-441 cell is sufficient to cleave the lambda repressor, under conditions where a normal basal level of RecA430 protein is also present allowing the formation of mixed multimers on single-stranded DNA regions normally present in the cell.
D010582:Bacteriophage lambda / Q000235:genetics*; D004277:DNA, Single-Stranded / Q000235:genetics / Q000378:metabolism; D005809:Genes, Regulator; D005816:Genetic Complementation Test; D008242:Lysogeny; D009154:Mutation; D011938:Rec A Recombinases / Q000235:genetics* / Q000378:metabolism; D014775:Virus Activation
D004277:DNA, Single-Stranded; D011938:Rec A Recombinases