KMID : 0617319940040010084
|
|
Journal of Pharmacetical Sceiences Ewha Womans University 1994 Volume.4 No. 1 p.84 ~ p.92
|
|
Alternative Pathways and Reactions of Benzyl Alcohol and Benzaldehyde with Horse Liver Alcohol Dehydrogenase
|
|
Shearer, Gretchen L.
Kim, Keehyuk/Lee, Kang Man/Wang, Kathy C./Plapp, Bryce V.
|
|
Abstract
|
|
|
Liver alcohol dehydragenase catalyzes the reaction of NAD^+ and benzyl alcohol to form NADH and benzaldehyde by a predominantly ordered reaction. However, enzyme-alcohol binary and abortive ternary complexes form at high concentrations of benzyl alcohol, and benzaldehyde is slowly oxidized to benzoic acid. Steady-state and transient kinetic studies, equilibrium spectrophotometric measurements, product analysis, and kinetic simulations provide estimates of rate constants for a complete mechanism with the following reactions: (1) E??E-NAD^+?? E-NAD^+-RCH_2OH ?? E-NADH-RCHO??E-NADH??E; (2) E-NADH??E-NADH-RCH_2OH??E-RCH_2OH??E; (3) E-NAD^+ ?? E-NAD^+-RCHO ¡æ E-NADH-RCOOH??E-NADH. The internal equilibrium constant for hydrogen transfer determined at 30¡É and pH7 is about 5:1 in favor of E-NAD^+-RCH_2OH and has a complex pH dependence. Benzyl alcohol binds weakly to free enzyme (K_d=7mM) and significantly decreases the rates of binding of NAD^+ and NADH. The reaction of NAD^+ and benzyl alcohol is therefore kinetically ordered, not random. High concentrations of benzyl alcohol (>1 mM) inhibit turnover by formation of the abortive E-NADH-RCH_2-OH complex, which dissociates at 0.3s^-1 as compared to 6.3s^-1 for E-NADH. The oxidation of benzaldehyde by E- NAD^+(K_m=15mM, V/E = 0.4 s^-1 ) is inefficient relative to the oxidation of benzyl alcohol (K_m=28 §, V/E = 3.1 s^-1) and leads to a dismutation (2RCHO ¡æ RCH_2OH + RCOOH) as E-NADH reduces benzaldehyde. The results provide a description of final product distributions for the alternative reactions catalyzed by the multifunctional enzyme.
|
|
KEYWORD
|
|
|
|
FullTexts / Linksout information
|
|
|
|
Listed journal information
|
|
|
|