The Bacillus subtilis genome comprises two paralogous single-stranded DNA binding protein (SSB) genes, ssb and ywpH, which show distinct expression patterns. The main ssb gene is strongly expressed during exponential growth and is coregulated with genes encoding the ribosomal proteins S6 and S18. The gene organization rpsF-ssb-rpsR as observed in B. subtilis is found in many gram-positive as well as some gram-negative bacteria, but not in Escherichia coli. The ssb gene is essential for cell viability, and like other SSBs its expression is elevated during SOS response. In contrast, the paralogous ywpH gene is transcribed from its own promoter at the onset of stationary phase in minimal medium only. Its expression is ComK dependent and its gene product is required for optimal natural transformation.Single-stranded DNA binding proteins (SSBs) in bacteria play crucial roles in DNA replication, repair, and recombination processes. The function of SSB in these processes, its biochemical properties, and its interaction with other proteins in the cell have been studied extensively in Escherichia coli and several bacteriophages (17,26). Relatively little is known about the regulation of SSB expression.In Escherichia coli the ssb gene is preceded by three promoters, one of them being inducible by DNA damage (3, 4). The DNA damage inducibility is due to the presence of a LexA binding site in the upstreammost promoter. Interestingly, this SOS-box is shared with the divergently transcribed uvrA gene, coding for the A subunit of the exonuclease ABC, which is involved in DNA repair (3). The same organization was found for the uvrA and ssb genes in Sinorhizobium meliloti (24). Although this gene organization is also identical in Proteus mirabilis and Serratia marcescens, the ssb genes of these bacteria are not inducible by DNA damage (7). It has been suggested that E. coli SSB negatively autoregulates its own translation, because it is capable of binding to its own mRNA, in this way inhibiting translation (21).Two paralogous genes coding for SSB were found in the Bacillus subtilis genome, ssb and ywpH. The deduced amino acid sequences of SSB and YwpH show 80% similarity and 63% identity. Notably, YwpH is lacking 66 amino acid residues of the C terminus of SSB. Although the amino acid sequences of bacterial SSBs are highly conserved within the first two thirds of the protein containing the DNA-binding domains, they diverge substantially in the C-terminal third region (7). The C-terminal region of E. coli SSB is not required for DNA binding in vitro, but is essential for its in vivo function (6, 29). In contrast to E. coli, neither of the B. subtilis ssb genes is organized adjacent to uvrA as the ssb gene is in E. coli. The first one, ssb, maps at 358.6 o of the B. subtilis genome and is flanked by the rpsF and rpsR genes, coding for the ribosomal proteins S6 and S18, respectively (Fig. 1A). A rho-independent transcriptional terminator is situated downstream of the rpsR gene, and possibly rpsF, ssb, and rpsR belong to one operon....

Bacillus coagulans has good potential as an industrial production organism for platform chemicals from renewable resources but has limited genetic tools available. Here, we present a targeted gene disruption system using the Cre-lox system, development of a LacZ reporter assay for monitoring gene transcription, and heterologous d-lactate dehydrogenase expression.

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The genes for DNA uptake and recombination in Bacilli are commonly regulated by the transcriptional factor ComK. We have identified a ComK homologue in Bacillus coagulans, an industrial relevant organism that is recalcitrant for transformation. Introduction of B. coagulans comK gene under its own promoter region into Bacillus subtilis comK strain results in low transcriptional induction of the late competence gene comGA, but lacking bistable expression. The promoter regions of B. coagulans comK and the comGA genes are recognized in B. subtilis and expression from these promoters is activated by B. subtilis ComK. Purified ComK protein of B. coagulans showed DNA-binding ability in gel retardation assays with B. subtilis-and B. coagulans-derived probes. These experiments suggest that the function of B. coagulans ComK is similar to that of ComK of B. subtilis. When its own comK is overexpressed in B. coagulans the comGA gene expression increases 40-fold, while the expression of another late competence gene, comC is not elevated and no reproducible DNAuptake could be observed under these conditions. Our results demonstrate that B. coagulans ComK can recognize several B. subtilis comK-responsive elements, and vice versa, but indicate that the activation of the transcription of complete sets of genes coding for a putative DNA uptake apparatus in B. coagulans might differ from that of B. subtilis.