Nickel-specific, slow-binding inhibition of carbon monoxide dehydrogenase from Rhodospirillum rubrum by cyanide.
The inhibition of purified carbon monoxide dehydrogenase from Rhodospirillum rubrum by cyanide was investigated in both the presence and absence of CO and electron acceptor. The inhibition was a time-dependent process exhibiting pseudo-first-order kinetics under both sets of conditions. The true second-order rate constants for inhibition were 72.2 M-1 s-1 with both substrates present and 48.9 and 79.5 M-1 s-1, respectively, for the reduced and oxidized enzymes incubated with cyanide. CO partially protected the enzyme against inhibition after 25-min incubation with 100 microM KCN. Dissociation constants of 8.46 microM (KCN) and 4.70 microM (CO) were calculated for the binding of cyanide and CO to the enzyme. Cyanide inhibition was fully reversible under an atmosphere of CO after removal of unbound cyanide. N2 was unable to reverse the inhibition. The competence of nickel-deficient (apo) CO dehydrogenase to undergo activation by NiCl2 was unaffected by prior incubation with cyanide. Cyanide inhibition of holo-CO dehydrogenase was not reversed by addition of NiCl2. 14CN- remained associated with holoenzyme but not with apoenzyme through gel filtration chromatography. These findings suggest that cyanide is a slow-binding, active-site-directed, nickel-specific, reversible inhibitor of CO dehydrogenase. We propose that cyanide inhibits CO dehydrogenase by being an analogue of CO and by binding through enzyme-bound nickel.
Ensign
S A
SA
Department of Biochemistry, College of Agricultural and Life Sciences, University of Wisconsin-Madison 53706.
Hyman
M R
MR
Ludden
P W
PW
D000445:Aldehyde Oxidoreductases / Q000037:antagonists & inhibitors*; D001665:Binding Sites / Q000187:drug effects; D001667:Binding, Competitive; D002248:Carbon Monoxide / Q000494:pharmacology; D003486:Cyanides / Q000494:pharmacology*; D004357:Drug Synergism; D004789:Enzyme Activation / Q000187:drug effects; D007700:Kinetics; D009097:Multienzyme Complexes; D009532:Nickel / Q000494:pharmacology*; D011190:Potassium Cyanide / Q000494:pharmacology*; D012247:Rhodospirillum rubrum / Q000201:enzymology*
D003486:Cyanides; D009097:Multienzyme Complexes; D009532:Nickel; D002248:Carbon Monoxide; D000445:Aldehyde Oxidoreductases; C019823:carbon monoxide dehydrogenase; D011190:Potassium Cyanide