OVERPRODUCTION OF THE HSD SUBUNITS LEADS TO THE LOSS OF TEMPERATURE-SENSITIVE RESTRICTION AND MODIFICATION PHENOTYPE

The genes hsdM and hsdS for M. EcoKI modification methyltrasferase and the complete set of hsdR, hsdM and hsdS genes coding for R. EcoKI res triction endonuclease, both with and without a temperature-sensitive ( ts) mutation in hrdS gene, were cloned in pBR322 plasmid and introduce d into E. coli C (a strain without a natural restriction-modification (R-M) system). The strains producing only the methyltransferase, or to gether with the endonuclease, were thus obtained. The hsdS(ts-1) mutat ion, mapped previously in the distal variable region of the hsdS gene with C-1245-T transition has no effect on the R-M phenotype expressed from cloned genes in bacteria grown at 42 degrees C. In clones transfo rmed with the whole hsd region an alleviation of R-M functions was obs erved immediately after the transformation, but after subculture the t ransformants expressed the wild-type R-M phenotype irrespective of whe ther the wild-type or the mutant hsdS allele was present in the hybrid plasmid. Simultaneous overproduction of HsdM and HsdM subunits impair s the ts effect of the hsdS(ts-1) mutation on restriction and modifica tion.