Redox titrations of carbon monoxide dehydrogenase from Clostridium thermoaceticum.
Redox titrations of carbon monoxide dehydrogenase (CODH) from Clostridium thermoaceticum were performed using the reductant CO and the oxidant thionin. Titrations were followed at 420 nm, a wavelength sensitive to redox changes of the iron-sulfur clusters in the enzyme. When CODH was oxidized by just enough thionin to maximize A420, two molecules of CO per mole of CODH dimer (4 equiv/mol) reduced the enzyme fully. Likewise, 4 equiv/mol of thionin oxidized the fully-reduced enzyme to the point where A420 maximized. The four n = 1 redox sites which titrated in this region were designated group I sites. They include at least two iron-sulfur clusters, [Fe/S]A and [Fe/S]B, and two other sites, A' and B'. The [Fe4S4]2+/1+ cluster in CODH is included in this group. [Fe/S]B and B' have reduction potentials (at pH 8) below -480 mV vs NHE; [Fe/S]A and A' have reduction potentials above that value. The reduction potential of either [Fe/S]B or B' is near to the CO/CO2 couple at pH 8 (-622 mV). When CODH was oxidized by more than enough thionin to maximize A420, some of the excess thionin oxidized the so-called group II redox sites. These sites have reduction potentials more positive than group I and do not exhibit changes at 420 nm when titrated. Titration of group II sites required 1-2 equiv/mol. EPR of reduced group II sites exhibited the gav = 1.82 signal. When these sites were oxidized, the only signal present had g values at 2.075, 2.036, and 1.983.(ABSTRACT TRUNCATED AT 250 WORDS)
Shin
W
W
Department of Chemistry, Texas A&M University, College Station 77843.
Stafford
P R
PR
Lindahl
P A
PA
D000445:Aldehyde Oxidoreductases / Q000378:metabolism*; D001665:Binding Sites; D002248:Carbon Monoxide / Q000378:metabolism; D003013:Clostridium / Q000201:enzymology*; D004578:Electron Spin Resonance Spectroscopy; D004789:Enzyme Activation / Q000187:drug effects; D007506:Iron-Sulfur Proteins / Q000378:metabolism; D046911:Macromolecular Substances; D009097:Multienzyme Complexes; D010084:Oxidation-Reduction; D010640:Phenothiazines / Q000378:metabolism / Q000494:pharmacology; D013053:Spectrophotometry
D007506:Iron-Sulfur Proteins; D046911:Macromolecular Substances; D009097:Multienzyme Complexes; D010640:Phenothiazines; D002248:Carbon Monoxide; D000445:Aldehyde Oxidoreductases; C019823:carbon monoxide dehydrogenase; C009469:thionine