IDENTIFICATION OF THE HISTIDINE PROTEIN-KINASE KINB IN PSEUDOMONAS-AERUGINOSA AND ITS PHOSPHORYLATION OF THE ALGINATE REGULATOR ALGB

The exopolysaccharide alginate is an important virulence factor in chr onic lung infections caused by the bacterium Pseudomonas aeruginosa. T wo positive activators for alginate synthesis, algB and algR, are memb ers of a superfamily of response regulators of the two-component regul atory system. AlgB belongs to the NtrC subfamily of response regulator s and is required for high-level production of alginate. In this study , an open reading frame encoding a polypeptide of 66 kh)a, designated kinB, was identified immediately downstream of algB. The sequence of K inB is homologous to the histidine protein kinase members of two-compo nent regulatory systems. Western blot analysis of a P. aeruginosa stra in carrying a kinB-lacZ protein fusion and studies of KinB-phoA fusion s indicate that KinB localizes to the inner membrane and has a NH2-ter minal periplasmic domain, A KinB derivative containing the COOH termin us of KinB was generated and purified. In the presence of [gamma-P-32] ATP, the purified COOH-terminal KinB protein was observed to undergo p rogressive autophosphorylation in vitro. Moreover, the phosphoryl labe l of KinB could be rapidly transferred to purified AlgB, Substitutions of the residues conserved among histidine protein kinases abolished K inB autophosphorylation, These results provide evidence that KinB enco des the AlgB cognate histidine protein kinase.