Coordination of selenium to molybdenum in formate dehydrogenase H from Escherichia coli.
Formate dehydrogenase H from Escherichia coli contains multiple redox centers, which include a molybdopterin cofactor, an iron-sulfur center, and a selenocysteine residue (SeCys-140 in the polypeptide chain) that is essential for catalytic activity. Here we show that addition of formate to the native enzyme induces a signal typical of Mo(V) species. This signal is detected by electron paramagnetic resonance (EPR) spectroscopy. Substitution of 77Se for natural isotope abundance Se leads to transformation of this signal, indicating a direct coordination of Se with Mo. Mutant enzyme with cysteine substituted at position 140 for the selenocysteine residue has decreased catalytic activity and exhibits a different EPR signal. Since determination of the Se content of wild-type enzyme indicates approximately 1 gram atom per mol, we conclude that it is the Se atom of the SeCys-140 residue in the protein that is coordinated directly with Mo. The amino acid sequence flanking the selenocysteine residue in formate dehydrogenase H is similar to a conserved sequence found in several other prokaryotic molybdopterin-dependent enzymes. In most of these other enzymes a cysteine residue, or in a few cases a serine or a selenocysteine residue, occurs in the position corresponding to SeCys-140 of formate dehydrogenase H. By analogy with formate dehydrogenase H in these other enzymes, at least one of the ligands to Mo should be provided by an amino acid residue of the protein. This ligand could be the Se of a selenocysteine residue, sulfur of a cysteine residue, or, in the case of a serine residue, oxygen.
Gladyshev
V N
VN
Laboratory of Biochemistry, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, MD 20892.
Khangulov
S V
SV
Axley
M J
MJ
Stadtman
T C
TC
D000595:Amino Acid Sequence; D017124:Conserved Sequence; D003545:Cysteine / Q000737:chemistry; D004578:Electron Spin Resonance Spectroscopy; D004926:Escherichia coli / Q000201:enzymology*; D005560:Formate Dehydrogenases / Q000737:chemistry* / Q000235:genetics; D006864:Hydrogenase / Q000737:chemistry* / Q000235:genetics; D007553:Isotope Labeling; D008024:Ligands; D008667:Metalloproteins / Q000737:chemistry* / Q000235:genetics; D008969:Molecular Sequence Data; D008982:Molybdenum / Q000737:chemistry*; D009097:Multienzyme Complexes / Q000737:chemistry* / Q000235:genetics; D012643:Selenium / Q000737:chemistry*; D017279:Selenocysteine / Q000737:chemistry; D017386:Sequence Homology, Amino Acid; D012694:Serine / Q000737:chemistry
D008024:Ligands; D008667:Metalloproteins; D009097:Multienzyme Complexes; D017279:Selenocysteine; D012694:Serine; D008982:Molybdenum; D006864:Hydrogenase; D005560:Formate Dehydrogenases; C054017:formate hydrogenlyase; D012643:Selenium; D003545:Cysteine