Pre-steady-state kinetics of the reactions of [NiFe]-hydrogenase from Chromatium vinosum with H2 and CO.

Publisher: Blackwell Science Ltd. on behalf of the Federation of European Biochemical Societies Country of Publication: England NLM ID: 0107600 Publication Model: Print Cited Medium: Print ISSN: 0014-2956 (Print) Linking ISSN: 00142956 NLM ISO Abbreviation: Eur J Biochem Subsets: MEDLINE

Publication: -2004: Oxford, UK : Blackwell Science Ltd. on behalf of the Federation of European Biochemical Societies
Original Publication: Berlin, New York, Springer.

Carbon Monoxide/*chemistry ; Chromatium/*enzymology ; Hydrogen/*chemistry ; Hydrogenase/*chemistry; Bacterial Proteins/chemistry ; Benzyl Viologen/chemistry ; Binding Sites ; Electron Spin Resonance Spectroscopy ; Oxidation-Reduction ; Oxygen/chemistry ; Paraquat/chemistry

Results are presented of the first rapid-mixing/rapid-freezing studies with a [NiFe]-hydrogenase. The enzyme from Chromatium vinosum was used. In particular the reactions of active enzyme with H2 and CO were monitored. The conversion from fully reduced, active hydrogenase (Nia-SR state) to the Nia-C* state was completed in less than 8 ms, a rate consistent with the H2-evolution activity of the enzyme. The reaction of CO with fully reduced enzyme was followed from 8 to 200 ms. The Nia-SR state did not react with CO. It was discovered, contrary to expectations, that the Nia-C* state did not react with CO when reactions were performed in the dark. When H2 was replaced by CO, a Nia-C* EPR signal appeared within 11 ms; this was also the case when H2 was replaced by Ar. With CO, however, the Nia-C* state decayed within 40 ms, due to the generation of the Nia-S.CO state (the EPR-silent state of the enzyme with bound CO). The Nia-C* state, induced after 11 ms by replacing H2 by CO in the dark, could be converted, in the frozen enzyme, into the EPR-detectable state with CO bound to nickel (Nia*.CO) by illumination at 30 K (evoking the Nia-L* state), followed by dark adaptation at 200 K. This can be explained by assuming that the Nia-C* state represents a formally trivalent state of nickel, which is unable to bind CO, whereas nickel in the Nia-L* and the Nia*.CO states is formally monovalent.

0 (Bacterial Proteins)
13096-46-3 (Benzyl Viologen)
7U1EE4V452 (Carbon Monoxide)
7YNJ3PO35Z (Hydrogen)
EC 1.12.- (nickel-iron hydrogenase)
EC 1.12.7.2 (Hydrogenase)
PLG39H7695 (Paraquat)
S88TT14065 (Oxygen)

Date Created: 19990327 Date Completed: 19990422 Latest Revision: 20190620

20250114

10.1046/j.1432-1327.1999.00057.x

10092843