Purification and Characterization of a Tungsten-Containing Formate Dehydrogenase from Desulfovibrio gigas

An air-stable formate dehydrogenase (FDH), an enzyme that catalyzes the oxidation of formate to carbon dioxide, was purified from the sulfate reducing organism Desulfovibrio gigas (D. gigas) NCIB 9332. D. gigas FDH is a heterodimeric protein [α (92 kDa) and β (29 kDa) subunits] and contains 7 ± 1 Fe...

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Bibliographic Details
Published in:Biochemistry (Easton) 1999-12, Vol.38 (49), p.16366-16372
Main Authors: Almendra, Maria J, Brondino, Carlos D, Gavel, Olga, Pereira, Alice S, Tavares, Pedro, Bursakov, Sergey, Duarte, Rui, Caldeira, Jorge, Moura, Jose J. G, Moura, Isabel
Format: Article
Language:English
Subjects:
Aerobiosis
Chromatography, Gel
Desulfovibrio - enzymology
Desulfovibrio gigas
Electron Spin Resonance Spectroscopy
Formate Dehydrogenases - chemistry
Formate Dehydrogenases - isolation & purification
Formate Dehydrogenases - metabolism
formic acid
Guanine Nucleotides - analysis
Metalloproteins - chemistry
Metalloproteins - isolation & purification
Metalloproteins - metabolism
Molecular Weight
Pterins - analysis
Spectrophotometry, Ultraviolet
Spectroscopy, Mossbauer
tungsten
Tungsten - chemistry
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Summary:An air-stable formate dehydrogenase (FDH), an enzyme that catalyzes the oxidation of formate to carbon dioxide, was purified from the sulfate reducing organism Desulfovibrio gigas (D. gigas) NCIB 9332. D. gigas FDH is a heterodimeric protein [α (92 kDa) and β (29 kDa) subunits] and contains 7 ± 1 Fe/protein and 0.9 ± 0.1 W/protein. Selenium was not detected. The UV/visible absorption spectrum of D. gigas FDH is typical of an iron−sulfur protein. Analysis of pterin nucleotides yielded a content of 1.3 ± 0.1 guanine monophosphate/mol of enzyme, which suggests a tungsten coordination with two molybdopterin guanine dinucleotide cofactors. Both Mössbauer spectroscopy performed on D. gigas FDH grown in a medium enriched with 57Fe and EPR studies performed in the native and fully reduced state of the protein confirmed the presence of two [4Fe-4S] clusters. Variable-temperature EPR studies showed the presence of two signals compatible with an atom in a d1 configuration albeit with an unusual relaxation behavior as compared to the one generally observed for W(V) ions.
ISSN:0006-2960
1520-4995
DOI:10.1021/bi990069n