STUDIES ON THE HETEROLOGOUS EXPRESSION OF BSTVI RESTRICTION-ENDONUCLEASE IN ESCHERICHIA-COLI

Bacterial restriction and modification systems must be regulated to av oid self-restriction It is generally accepted that cognate DNA methylt ransferases normally protects both, the host's chromosome and extrachr omosomal elements from the activity of their endonuclease counterparts . When the bstVIRM genes from Bacillus stearothermophilusV were subclo ned into Escherichia coli, several clones exhibiting a r(+)m(-) phenot ype were originated. The present work was undertaken to analyze the po ssibility that mechanisms other than DNA methylation could account for the viability of these cells. No evidence was found for an inhibitory agent or endonuclease compartmentation. In vivo experiments showed th at lambda phage multiplication was poorly restricted by the heterologo us enzyme. The restricting activity against the incoming phage increas ed however when phage adsortion was performed at higher temperatures. Analogous experiments in which a DNA-repair deficient strain was used as a host for the thermophilic R-M system suggested, to some extent, t he participation of the repair machinery in the viability of r(+)m(-) clones.