THE BACILLUS-SUBTILIS CRH GENE ENCODES A HPR-LIKE PROTEIN INVOLVED INCARBON CATABOLITE REPRESSION

Carbon catabolite repression (CCR) of several Bacillus subtilis catabo lic genes is mediated by ATP-dependent phosphorylation of histidine-co ntaining protein (HPr), a phosphocarrier protein of the phosphoenolpyr uvate (PEP): sugar phosphotransferase system, In this study, we report the discovery of a new B, subtilis gene encoding a HPr-like protein, Crh (for catabolite repression HPr), composed of 85 amino acids. Crh e xhibits 45% sequence identity with HPr, but the active site His-15 of HPr is replaced with a glutamine in Crh, Crh is therefore not phosphor ylated by PEP and enzyme I, but is phosphorylated by ATP and the HPr k inase in the presence of fructose-1,6-bisphosphate. We determined Ser- 46 as the site of phosphorylation in Crh by carrying out mass spectrom etry with peptides obtained by tryptic digestion or CNBr cleavage, In a B. subtilis pfsH1 mutant strain, synthesis of beta-xylosidase, inosi tol dehydrogenase, and levanase was only partially relieved from CCR, Additional disruption of the crh gene caused almost complete relief fr om CCR, In a ptsH1 crh1 mutant, producing HPr and Crh in which Ser-46 is replaced with a nonphosphorylatable alanyl residue, expression of b eta-xylosidase was also completely relieved from glucose repression, T hese results suggest that CCR of certain catabolic operons requires, i n addition to CcpA, ATP-dependent phosphorylation of Crh, and HPr at S er-46.