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Molecular Organization of Intrinsic Restriction and Modification Genes BsuM of Bacillus subtilis Marburg

Graduate School for Advanced Study, National Institute of Genetics, Mishima 411-8540,¹ Department of Biochemistry and Molecular Biology, Faculty of Science, Saitama University, Saitama 338-8570, Saitama, Japan²

Received 25 June 2001/ Accepted 5 October 2001

Transcriptional analysis and disruption of five open reading frames (ORFs), ydiO, ydiP, ydiR, ydiS, and ydjA, in the prophage 3 region of the chromosome of Bacillus subtilis Marburg revealed that they are component genes of the intrinsic BsuM restriction and modification system of this organism. The classical mutant strain RM125, which lacks the restriction and modification system of B. subtilis Marburg, lacks the prophage 3 region carrying these five ORFs. These ORFs constitute two operons, the ydiO-ydiP operon and the ydiR-ydiS-ydjA operon, both of which are expressed during the logarithmic phase of growth. The predicted gene products YdiO and YdiP are the orthologues of cytosine DNA methyltransferases. The predicted YdiS product is an orthologue of restriction nucleases, while the predicted YdiR and YdjA products have no apparent paralogues and orthologues whose functions are known. Disruption of the ydiR-ydiS-ydjA operon resulted in enhanced transformation by plasmid DNA carrying multiple BsuM target sequences. Disruption of ydiO or ydiP function requires disruption of at least one of the following genes on the chromosome: ydiR, ydiS, and ydjA. The degrees of methylation of the BsuM target sequences on chromosomal DNAs were estimated indirectly by determining the susceptibility to digestion with XhoI (an isoschizomer of BsuM) of DNAs extracted from the disruptant strains. Six XhoI (BsuM) sites were examined. XhoI digested at the XhoI sites in the DNAs from disruptants with disruptions in both operons, while XhoI did not digest at the XhoI sites in the DNAs from the wild-type strain or from the disruptants with disruptions in the ydiR-ydiS-ydjA operon. Therefore, the ydiO-ydiP operon and the ydiR-ydiS-ydjA operon are considered operons that are responsible for BsuM modification and BsuM restriction, respectively.

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