A NOVEL MULTICOMPONENT REGULATORY SYSTEM MEDIATES H-2 SENSING IN ALCALIGENES-EUTROPHUS

Oxidation of molecular hydrogen catalyzed by [NiFe] hydrogenases is a widespread mechanism of energy generation among prokaryotes. Biosynthe sis of the H-2-oxidizing enzymes is a complex process subject to posit ive control by H-2 and negative control by organic energy sources. In this report we describe a novel signal transduction system regulating hydrogenase gene (hox) expression in the proteobacterium Alcaligenes e utrophus. This multicomponent system consists of the proteins HoxB, Ho sC, HoxJ, and HoxA. HoxB and HoxC share characteristic features of di meric [NiFe] hydrogenases and form the putative H-2 receptor that inte racts directly or indirectly with the histidine protein kinase HoxJ. A single amino acid substitution (HoxJG422S) in a conserved C-termina l glycine-rich moth of HoxJ resulted in a loss of H-2-dependent signa l transduction and a concomitant block in autophosphorylating activity , suggesting that autokinase activity is essential for the response to H-2. Whereas deletions in hoxB or hoxC abolished hydrogenase synthesi s almost completely, the autokinase deficient strain maintained high-l evel hox gene expression, indicating that the active sensor kinase exe rts a negative effect on hox gene expression in the absence of H-2. Su bstitutions of the conserved phosphoryl acceptor residue Asp55 in the response regulator HoxA (HoxAD55E and HoxAD55N) disrupted the H-2 sign al-transduction chain. Unlike other NtrC-like regulators, the altered HoxA proteins still allowed high-level transcriptional activation. The data presented here suggest a model in which the nonphosphorylated fo rm of HoxA stimulates transcription in concert with a yet unknown glob al energy-responsive factor.