Isotopic analysis of the reaction catalyzed by glycerol dehydrogenase

Betty N. Leichus, John S. Blanchard

Research output: Contribution to journalArticle

5 Citations (Scopus)

Abstract

Glycerol dehydrogenase catalyzes the reversible NAD+-dependent oxidation of glycerol to form dihydroxyacetone. Initial velocity, product, and dead-end inhibition studies performed for the forward and reverse reactions support an ordered kinetic mechanism with NAD+ binding first and NADH released last. A monovalent cation is required for enzymatic activity and glycerol binding, with K+ having the highest activity as measured by V. The pH dependence of the kinetic parameters V and V/Kglycerol, as well as the temperature dependence of the VpH profile, suggested that an enzymic carboxylate group functions as a base in catalysis. The pH dependence of the primary deuterium kinetic isotope effect shows that DV/Kglycerol increases from a pH-independent value of 1.15 at high pH values to a pH-independent value of 2.44 at low pH values. DV exhibits a similar pH dependence, increasing from a pH-independent value of 2.57 at high pH values to a pH independent value of 4.88 at low pH values. A chemical mechanism for enzymatic glycerol oxidation is proposed based on the data.

Original languageEnglish (US)
Pages (from-to)14642-14649
Number of pages8
JournalBiochemistry®
Volume33
Issue number48
StatePublished - 1994

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glycerol dehydrogenase
NAD
Glycerol
Dihydroxyacetone
Monovalent Cations
Oxidation
Kinetics
Deuterium
Kinetic parameters
Isotopes
Catalysis
Temperature

ASJC Scopus subject areas

  • Biochemistry

Cite this

Isotopic analysis of the reaction catalyzed by glycerol dehydrogenase. / Leichus, Betty N.; Blanchard, John S.

In: Biochemistry®, Vol. 33, No. 48, 1994, p. 14642-14649.

Research output: Contribution to journalArticle

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