Characterization of the oxygen tolerance of a hydrogenase linked to a carbon monoxide oxidation pathway in Rubrivivax gelatinosus.
A hydrogenase linked to the carbon monoxide oxidation pathway in Rubrivivax gelatinosus displays tolerance to O2. When either whole-cell or membrane-free partially purified hydrogenase was stirred in full air (21% O2, 79% N2), its H2 evolution activity exhibited a half-life of 20 or 6 h, respectively, as determined by an anaerobic assay using reduced methyl viologen. When the partially purified hydrogenase was stirred in an atmosphere containing either 3.3 or 13% O2 for 15 min and evaluated by a hydrogen-deuterium (H-D) exchange assay, nearly 80 or 60% of its isotopic exchange rate was retained, respectively. When this enzyme suspension was subsequently returned to an anaerobic atmosphere, more than 90% of the H-D exchange activity was recovered, reflecting the reversibility of this hydrogenase toward O2 inactivation. Like most hydrogenases, the CO-linked hydrogenase was extremely sensitive to CO, with 50% inhibition occurring at 3.9 microM dissolved CO. Hydrogen production from the CO-linked hydrogenase was detected when ferredoxins of a prokaryotic source were the immediate electron mediator, provided they were photoreduced by spinach thylakoid membranes containing active water-splitting activity. Based on its appreciable tolerance to O2, potential applications of this hydrogenase are discussed.
Maness
Pin-Ching
PC
The National Renewable Energy Laboratory, Golden, Colorado 80401-3393, USA. pinching_maness@nrel.gov
Smolinski
Sharon
S
Dillon
Anne C
AC
Heben
Michael J
MJ
Weaver
Paul F
PF
D002248:Carbon Monoxide / Q000378:metabolism* / Q000494:pharmacology; D006859:Hydrogen / Q000378:metabolism; D006864:Hydrogenase / Q000378:metabolism*; D010084:Oxidation-Reduction; D010100:Oxygen / Q000378:metabolism*; D010782:Photolysis; D020560:Proteobacteria / Q000187:drug effects / Q000378:metabolism*; D014867:Water / Q000737:chemistry
D014867:Water; D002248:Carbon Monoxide; D006859:Hydrogen; D006864:Hydrogenase; D010100:Oxygen