Regulation of the Bacillus subtilis phosphotransacetylase gene

The enzyme, phosphotransacetylase (Pta), catalyzes the conversion of acetyl coenzyme A to acetyl phosphate, The putative pta gene of Bacillus subtilis , which had been sequenced as part of the Genome Project, was cloned and ov erexpressed in Escherichia coli, We confirmed that the gene encodes Pta by measuring the enzymatic activity of the purified protein. Insertional mutag enesis of the pta gene resulted in complete loss of the Pta activity, indic ating that B. subtilis contains, only one kind of pta gene, Expression of a pta-lacZ fusion was induced in the presence of excess glucose in the growt h medium, and the intact ccpA gene was required for this activation. The tr anscriptional start site of the pta gene was located at 37 nucleotides upst ream of the pta start codon, and a cre (catabolite responsive element) sequ ence, a cis-acting element that is responsible for the catabolite repressio n of a number of carbon utilization genes in B. subtilis, was identified up stream of the tentative promoter site. Experiments involving oligonucleotid e-directed mutagenesis showed that the cre sequence is involved in glucose- mediated transcriptional activation.