Alpha-secondary tritium kinetic isotope effects for the hydrolysis of alpha-D-glucopyranosyl fluoride by exo-alpha-glucanases.

H. Matsui, John S. Blanchard, Curtis F. Brewer, E. J. Hehre

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Abstract

alpha-Secondary tritium kinetic isotope effects ranging from 1.17 to 1.26 were measured for the hydrolysis of alpha-D-glucopyranosyl fluoride (forming beta-D-glucose) catalyzed by several glucoamylases and a glucodextranase. These results indicate that cleavage of the C-F bond is slow and that the enzymic transition state has significant oxo-carbonium ion character. Strong support for this conclusion is provided by the agreement found in the case of Rhizopus niveus glucoamylase (alpha-TV/K 1.26; Km 26 mM) between measured values of the alpha-secondary deuterium kinetic isotope effects (alpha-DV/K 1.16; alpha-DV 1.20) and those calculated from the tritium isotope effect. The data are consistent with the promotion of an intramolecular elimination of fluoride by the present exo-alpha-glucanases based on their ability to stabilize, perhaps with a counter ion, the development of a carbonium ion-like transition state. Although the oxo-carbonium ion is formally denoted as an intermediate it could represent a transition state along a reaction pathway to a covalent glucosyl intermediate.

Original languageEnglish (US)
Pages (from-to)8714-8716
Number of pages3
JournalJournal of Biological Chemistry
Volume264
Issue number15
StatePublished - May 25 1989

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Tritium
Isotopes
Glucan 1,4-alpha-Glucosidase
Hydrolysis
exo-1,6-alpha-glucosidase
Ions
Kinetics
Radiation counters
Deuterium
Fluorides
Rhizopus
Glucose
glucosyl fluoride

ASJC Scopus subject areas

  • Biochemistry

Cite this

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title = "Alpha-secondary tritium kinetic isotope effects for the hydrolysis of alpha-D-glucopyranosyl fluoride by exo-alpha-glucanases.",
abstract = "alpha-Secondary tritium kinetic isotope effects ranging from 1.17 to 1.26 were measured for the hydrolysis of alpha-D-glucopyranosyl fluoride (forming beta-D-glucose) catalyzed by several glucoamylases and a glucodextranase. These results indicate that cleavage of the C-F bond is slow and that the enzymic transition state has significant oxo-carbonium ion character. Strong support for this conclusion is provided by the agreement found in the case of Rhizopus niveus glucoamylase (alpha-TV/K 1.26; Km 26 mM) between measured values of the alpha-secondary deuterium kinetic isotope effects (alpha-DV/K 1.16; alpha-DV 1.20) and those calculated from the tritium isotope effect. The data are consistent with the promotion of an intramolecular elimination of fluoride by the present exo-alpha-glucanases based on their ability to stabilize, perhaps with a counter ion, the development of a carbonium ion-like transition state. Although the oxo-carbonium ion is formally denoted as an intermediate it could represent a transition state along a reaction pathway to a covalent glucosyl intermediate.",
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T1 - Alpha-secondary tritium kinetic isotope effects for the hydrolysis of alpha-D-glucopyranosyl fluoride by exo-alpha-glucanases.

AU - Matsui, H.

AU - Blanchard, John S.

AU - Brewer, Curtis F.

AU - Hehre, E. J.

PY - 1989/5/25

Y1 - 1989/5/25

N2 - alpha-Secondary tritium kinetic isotope effects ranging from 1.17 to 1.26 were measured for the hydrolysis of alpha-D-glucopyranosyl fluoride (forming beta-D-glucose) catalyzed by several glucoamylases and a glucodextranase. These results indicate that cleavage of the C-F bond is slow and that the enzymic transition state has significant oxo-carbonium ion character. Strong support for this conclusion is provided by the agreement found in the case of Rhizopus niveus glucoamylase (alpha-TV/K 1.26; Km 26 mM) between measured values of the alpha-secondary deuterium kinetic isotope effects (alpha-DV/K 1.16; alpha-DV 1.20) and those calculated from the tritium isotope effect. The data are consistent with the promotion of an intramolecular elimination of fluoride by the present exo-alpha-glucanases based on their ability to stabilize, perhaps with a counter ion, the development of a carbonium ion-like transition state. Although the oxo-carbonium ion is formally denoted as an intermediate it could represent a transition state along a reaction pathway to a covalent glucosyl intermediate.

AB - alpha-Secondary tritium kinetic isotope effects ranging from 1.17 to 1.26 were measured for the hydrolysis of alpha-D-glucopyranosyl fluoride (forming beta-D-glucose) catalyzed by several glucoamylases and a glucodextranase. These results indicate that cleavage of the C-F bond is slow and that the enzymic transition state has significant oxo-carbonium ion character. Strong support for this conclusion is provided by the agreement found in the case of Rhizopus niveus glucoamylase (alpha-TV/K 1.26; Km 26 mM) between measured values of the alpha-secondary deuterium kinetic isotope effects (alpha-DV/K 1.16; alpha-DV 1.20) and those calculated from the tritium isotope effect. The data are consistent with the promotion of an intramolecular elimination of fluoride by the present exo-alpha-glucanases based on their ability to stabilize, perhaps with a counter ion, the development of a carbonium ion-like transition state. Although the oxo-carbonium ion is formally denoted as an intermediate it could represent a transition state along a reaction pathway to a covalent glucosyl intermediate.

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