Achromobacter, Alcaligenes and Related Genera

Abstract

The tetrameric cytoplasmic [NiFe] hydrogenase (SH) of Ralstonia eutropha couples the oxidation of hydrogen to the reduction of NAD under aerobic conditions. In the catalytic subunit HoxH, all six conserved motifs surrounding the [NiFe] site are present. Five of these motifs were altered by site-directed mutagenesis in order to dissect the molecular mechanism of hydrogen activation. Based on phenotypic characterizations, 27 mutants were grouped into four different classes. Mutants of the major class, class I, failed to grow on hydrogen and were devoid of H2 mutants revealed a high D2/H H16L) displayed D2/H -oxidizing activity. In one of these isolates (HoxH I64A), H2 exchange rate relative to a low H2 -sensitive growth on hydrogen due to an O2 binding was impaired. Class II -oxidizing activity. A representative (HoxH affected in class III mutants. Mutants forming class IV showed a particularly interesting phenotype. They displayed O2 exchange but had lost electron acceptor-reducing activity. Both activities were equally -sensitive SH protein.