Rate constants for a mechanism including intermediates in the interconversion of ternary complexes by horse liver alcohol dehydrogenase
- Univ. of Iowa, Iowa City (USA)
Transient kinetic data for partial reactions of alcohol dehydrogenase and simulations of progress curves have led to estimates of rate constants for the following mechanism, at pH 8.0 and 25{degrees}C: E in equilibrium E-NAD+ in equilibrium *E-NAD+ in equilibrium E-NAD(+)-RCH2OH in equilibrium E-NAD+-RCH2O- in equilibrium *E-NADH-RCHO in equilibrium E-NADH-RCHO in equilibrium E-NADH in equilibrium E. Previous results show that the E-NAD+ complex isomerizes with a forward rate constant of 620 s-1. The enzyme-NAD(+)-alcohol complex has a pK value of 7.2 and loses a proton rapidly (greater than 1000 s-1). The transient oxidation of ethanol is 2-fold faster in D{sub 2}O, and proton inventory results suggest that the transition state has a charge of -0.3 on the substrate oxygen. Rate constants for hydride ion transfer in the forward or reverse reactions were similar for short-chain aliphatic substrates (400-600 s-1). A small deuterium isotope effect for transient oxidation of longer chain alcohols is apparently due to the isomerization of the E-NAD+ complex. The transient reduction of aliphatic aldehydes showed no primary deuterium isotope effect; thus, an isomerization of the E-NADH-aldehyde complex is postulated, as isomerization of the E-NADH complex was too fast to be detected. The estimated microscopic rate constants show that the observed transient reactions are controlled by multiple steps.
- OSTI ID:
- 7063980
- Journal Information:
- Biochemistry; (USA), Vol. 29:18; ISSN 0006-2960
- Country of Publication:
- United States
- Language:
- English
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Related Subjects
ALCOHOL DEHYDROGENASE
BIOCHEMICAL REACTION KINETICS
DEUTERIUM
ISOTOPE EFFECTS
HORSES
ISOMERIZATION
LIVER
MATHEMATICAL MODELS
PROTONS
SPECIFICITY
SUBSTRATES
TRACER TECHNIQUES
ANIMALS
BARYONS
BODY
CHEMICAL REACTIONS
DIGESTIVE SYSTEM
ELEMENTARY PARTICLES
ENZYMES
FERMIONS
GLANDS
HADRONS
HEMIACETAL DEHYDROGENASES
HYDROGEN ISOTOPES
ISOTOPE APPLICATIONS
ISOTOPES
KINETICS
LIGHT NUCLEI
MAMMALS
NUCLEI
NUCLEONS
ODD-ODD NUCLEI
ORGANS
OXIDOREDUCTASES
REACTION KINETICS
STABLE ISOTOPES
VERTEBRATES
550201* - Biochemistry- Tracer Techniques