The mechanism of the quinone reductase reaction of pig heart lipoamide dehydrogenase.
The relationship between the NADH:lipoamide reductase and NADH:quinone reductase reactions of pig heart lipoamide dehydrogenase (EC 1.6.4.3) was investigated. At pH 7.0 the catalytic constant of the quinone reductase reaction (kcat.) is 70 s-1 and the rate constant of the active-centre reduction by NADH (kcat./Km) is 9.2 x 10(5) M-1.s-1. These constants are almost an order lower than those for the lipoamide reductase reaction. The maximal quinone reductase activity is observed at pH 6.0-5.5. The use of [4(S)-2H]NADH as substrate decreases kcat./Km for the lipoamide reductase reaction and both kcat. and kcat./Km for the quinone reductase reaction. The kcat./Km values for quinones in this case are decreased 1.85-3.0-fold. NAD+ is a more effective inhibitor in the quinone reductase reaction than in the lipoamide reductase reaction. The pattern of inhibition reflects the shift of the reaction equilibrium. Various forms of the four-electron-reduced enzyme are believed to reduce quinones. Simple and 'hybrid ping-pong' mechanisms of this reaction are discussed. The logarithms of kcat./Km for quinones are hyperbolically dependent on their single-electron reduction potentials (E1(7]. A three-step mechanism for a mixed one-electron and two-electron reduction of quinones by lipoamide dehydrogenase is proposed.
Vienozinskis
J
J
Institute of Biochemistry, Lithuanian Academy of Sciences, Vilnius, U.S.S.R.
Butkus
A
A
Cenas
N
N
Kulys
J
J
D000818:Animals; D016227:Benzoquinones; D008058:Dihydrolipoamide Dehydrogenase / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D009206:Myocardium / Q000201:enzymology*; D009243:NAD / Q000378:metabolism; D010084:Oxidation-Reduction; D011808:Quinone Reductases / Q000378:metabolism*; D011809:Quinones / Q000378:metabolism; D013552:Swine
D016227:Benzoquinones; D011809:Quinones; D009243:NAD; C004532:quinone; D011808:Quinone Reductases; D008058:Dihydrolipoamide Dehydrogenase