THE HUPTUV OPERON IS INVOLVED IN NEGATIVE CONTROL OF HYDROGENASE SYNTHESIS IN RHODOBACTER-CAPSULATUS

The hupT, hupU, and hupV genes, which are located upstream from the hu pSLC and hypF genes in the chromosome of Rhodobacter capsulatus, form the hupTUV operon expressed from the hupT promoter. The hupU and hupV genes, previously thought to belong to a single open reading frame, en code HupU, of 34.5 kDa (332 amino acids), and HupV, of 50.4 kDa (476 a mino acids) which are greater than or equal to 50% identical to the ho mologous Bradyrhizobium japonicum HupU and HupV proteins and Rhodobact er sphaeroides HupU1 and HupU2 proteins, respectively; they also have 20 and 29% similarity with the small subunit (HupS) and the large subu nit (HupL), respectively, of R. capsulatus [NiFe]hydrogenase. HupU lac ks the signal peptide of HupS and HupV lacks the C-terminal sequence o f HupL, which are cleaved during hydrogenase processing. Inactivation of hupV by insertional mutagenesis or of hupUV by in-frame deletion le d to HupV(-) and Hup(UV)(-) mutants derepressed by hydrogenase synthes is, particularly in the presence of oxygen. These mutants were complem ented in trans by plasmid-borne hupTUV but not by hupT or by hupUV, ex cept when expressed from the inducible fur promoter. Complementation o f the HupV(-) and Hup(UV)(-) mutants brought about a decrease in hydro genase activity up to 10-fold, to the level of the wild-type strain B1 0, indicating that HupU and HupV participate in negative regulation of hydrogenase expression in concert with HupT, a sensor histidine kinas e involved in the repression process. Plasmid-borne gene fusions used to monitor hupTUV expression indicated that the operon is expressed at a low level (50- to 100-fold lower than hupS).