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Abstract

Using a PCR-based approach, we have cloned various a factor homologous genes from Clostridium acetobutylicum DSM 792. The nucleotide sequence of the dnaE-sigA operon has been determined and predicts two genes encoding 69- and 43-kDa proteins. The deduced DnaE amino acid sequence has approximately 30% amino acid identity with protein sequences of other primases. The putative sigA gene product shows high homology to primary a factors of various bacteria, most significantly to Bacillus subtilis and Staphylococcus aureus. Northern (RNA) blot analysis revealed that both genes form an operon, which is clearly expressed under conditions that allow for cell division. A promoter sequence with significant homology to the sigma(H)-dependent Bacillus promoters preceded the determined transcriptional start point, 182 bp upstream of the GUG start codon of dnaE. The homologous genes to Bacillus spp. sporulation sigma factors G, E, and K have been cloned and sequenced. Indirect evidence for the existence of sigma(F) was obtained by identification of a DNA sequence homologous to the respective Bacillus consensus promoter. Southern hybridization analysis indicated the presence of sigma(D) and sigma(H) homologous genes in C. acetobutylicum. A new gene group conserved within the eubacteria, but with yet unspecified functions, is described. The data presented here provide strong evidence that at least some of the complex regulation features of sporulation in B. subtilis are conserved in C. acetobutylicum and possibly Clostridium spp.