Spectroscopic and kinetic characterization of active site mutants of Desulfovibriofructosovorans Ni-Fe hydrogenase

Site-directed mutagenesis of amino acid residues proximate to the active site of the Ni-Fe hydrogenase of Desulfovibrio fructosovorans has been done. The different mutants have been analyzed by FTIR spectroscopy and compared with wild type enzyme. The changes observed in the spectra confirm that hydrogen bonds between the CN– ligands of the active site's Fe atom and certain neighbor amino acid residues stabilize the active center within the protein matrix. However, kinetic analysis of the mutants indicates that none of the replaced residues have an important role in the catalytic mechanism of the hydrogenase. Electronic Supplementary Material is available if you access this article at http://dx.doi.org/10.1007/s00775-002-0397-4. On that page (frame on the left side), a link takes you directly to the supplementary material.