In Azotobacter chroococcum the hydrogenase gene (hup) cluster spans.about 14 kb of DNA. The genes coding for the small and large subunits, hupSL, are located at the 5' end, and a cluster of genes, hupABYCDE, resembling the Escherichia coli hyp operon, is located at the 3' end. In this study, we determined the effect of adding nickel to the medium used for the growth of hup mutants. Hydrogenase activity was restored to hupA and hupB mutants, but not hup Y, hupD, or hupE mutants, by the addition of nickel to the growth medium, suggesting that the products of hupA and hupB are somehow involved in nickel metabolism. The restoration of hydrogenase activity to the hupB mutant required protein synthesis.