Cloning and sequencing of the genes encoding the subunits of bidirectional hydrogenase of Anabaena variabilis IAM M58

Abstract

The enzymes directly involved in hydrogen metabolism in cyanobacteria are hydrogenases and nitrogenase. Functionally, two types of hydrogenase are known: uptake hydrogenase (Hup) and bidirectional hydrogenase (Hox). Distribution of hydrogenases among 15 filamentous cyanobacteria was studied by heterologous Southern hybridizations with hupL and hoxH probes from Anabaena PCC7120 and by direct assay of in vitro hydrogenase activities. All tested 15 strains showed hybridization with the hupL probe. With hoxH probe, hybridization bands are detected in 12 of them, but not in 3 of them. Moreover, the latter have no detectable in vitro Hox activity. The hupL and/or hoxH gene from Anabaena PCC7120 were inactivated. The extracts from the hoxH- strains were completely unable to perform Na2S2O4- and methyl viologen-dependent H2 evolution. H2 uptake with PMS by extracts from N2 fixing cells was undetectable in hupL-. The hupL- and hupL-hoxH- strains produced 4-7 times higher amounts of H2 than the wild-type strains. The nitrogenase activities of these hupL- strains were not reduced compared to the wild-type. Unexpectedly, the hoxH- strains were lower in H2 production activities by 20-50% compared to the wild-type.