GENETIC-CHARACTERISTICS OF NEW RECA MUTANTS OF ESCHERICHIA-COLI K-12

To search for functionally thermosensitive (FT) recA mutations, as wel l as mutations with differently affect RecA protein functions, seven n ew recA mutations in three different regions of the RecA protein struc ture proposed by Story et al. [R. M. Story, I. T. Weber, and T. A. Ste itz, Nature (London) 355:318-325, 1992] were constructed. Additionally , the recA2283 allele responsible for the FT phenotype of the recA200 mutant was sequenced. Five single mutations (recA2277, recA2278, recA2 283, recA2283E, and recA2284) and one double mutation (recA2278-5) gen erated, respectively, the amino acid substitutions L-277-->N, G-278--> P, L-283-->P, L-283-->E, I-284-->D, and G-278-->T plus V-275-->F in th e alpha-helix H-beta-strand 9 region of the C-terminal domain of the R ecA protein structure. According to recombination, repair, and SOS-ind ucible characteristics, these six mutations fall into four phenotypic classes: (i) an FT class, with either inhibition of all three analyzed functions at 42 degrees C (recA2283), preferable inhibition at 42 deg rees C of recombination and the SOS response (recA2278), or inhibition at 42 degrees C of only recombination (recA2278-5); (ii) a moderately deficient class (recA2277); (iii) a nondeficient class (recA2283E); a nd (iv) a mutation with a null phenotype (recA2284). The recA2223 muta tion generates an L-223-->M substitution in beta-strand 6 in a central domain of the RecA structure. This FT mutation shows preferable inhib ition of the SOS response at 42 degrees C. The recA2183 mutation produ ces a K-183-->M substitution in alpha-helix F of the same domain. The Lys-183 position in the Escherichia coli RecA protein was found among positions which are important for interfilament interaction (R. M. Sto ry, I. T. Weber, and T. A. Steitz, Nature (London) 355:318-325, 1992).