Cloning and expression of Bacillus subtilis phage DNA methyltransferase genes in Escherichia coli and B. subtilis.
The DNA methyltransferase (Mtase) genes of the temperate Bacillus subtilis phages SPR (wild type and various mutants), phi 3T, rho 11 and SP beta have been cloned and expressed in Escherichia coli and B. subtilis host-plasmid vector systems. Mtase activity has been quantitated in these clones by performing in vitro methylation assays of cell-free extracts. The four-phage Mtase genes differ in the amount of Mtase synthesized when transcribed from their genuine promoters. In B. subtilis as well as in E. coli the SPR Mtase is always produced in smaller amounts than the other phage Mtases. Expression levels of the SPR Mtase are dependent on the strength of the upstream vector promoter sequences. Overproduction of the SPR wild-type and mutant enzymes was achieved in E. coli (inducible expression) by fusions to the lambda pL or the tac promoter and in B. subtilis (constitutive expression) by means of the phage SP02 promoter.
D001412:Bacillus subtilis / Q000201:enzymology / Q000235:genetics*; D001435:Bacteriophages / Q000201:enzymology / Q000235:genetics*; D003001:Cloning, Molecular; D004248:DNA (Cytosine-5-)-Methyltransferases / Q000235:genetics*; D004262:DNA Restriction Enzymes; D004926:Escherichia coli / Q000235:genetics*; D005796:Genes; D005814:Genes, Viral; D009154:Mutation; D010957:Plasmids; D013728:Terminator Regions, Genetic; D014158:Transcription, Genetic
D004248:DNA (Cytosine-5-)-Methyltransferases; D004262:DNA Restriction Enzymes