Scope and mechanism of carbohydrase action. Stereospecific hydration of 2,6-anhydro-1-deoxy-D-gluco-hept-1-enitol catalyzed by α- and β-glucosidases and an inverting exo-α-glucanase

Edward J. Hehre, Curtis F. Brewer, Takao Uchiyama, Peter Schlesselmann, Jochen Lehmann

Research output: Contribution to journalArticle

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Abstract

D-Glucosylases of several types have been found to catalyze specific hydration and glycosyl transfer reactions (the former predominating) when incubated with 2,6-anhydro-1-deoxy-D-gluco-hept-1-enitol. α-Glucosidases from Candida tropicalis and from rice convert this substrate into 1-deoxy-α-D-gluco-heptulose. The β-glucosidase from sweet almonds and an exo-1,6-glucanase (glucodextranase) from Arthrobacter globiformis convert the substrate to 1-deoxy-β-D-gluco-heptulose. The anomeric form of the sugar was identified by following 1H NMR spectra of enzyme-enitol digests. The C-1 methyl group protons of the α and β anomers resonated at 1.45 and 1.40 ppm, respectively. Nonenzymatic anomerization of the β to the α form of the sugar was noted and, at equilibrium, 13C and hydroxyl 1H NMR spectra showed the presence of only the α anomer. The present work provides the first demonstration of the ability of an "inverting" glucanase to create specific anomeric configuration de novo from a substrate lacking α or β configuration. Indeed, the actions of glucodextranase on this enolic glycosyl donor lead not only to 1-deoxy-β-D-gluco-heptulose but also to a 1-deoxy-α-D-gluco-heptulosyl transfer product when enitol is the acceptor. These results reveal a catalytic mechanism that is able to convert a substrate to products of either configuration, depending on the type of acceptor. A mechanism is proposed for the stereospecific hydration of 2,6-anhydro-1-deoxy-D-gluco-hept-1-enitol by each enzyme, involving an incipient glycosyl carbonium ion and assuming the presence at the active site of two carboxyl groups arranged to account for catalysis of reactions with appropriate glycosidic substrates. These observations provide strong evidence for the functional flexibility of the catalytic groups of glycosylases.

Original languageEnglish (US)
Pages (from-to)3557-3564
Number of pages8
JournalBiochemistry
Volume19
Issue number15
StatePublished - 1980

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exo-1,6-alpha-glucosidase
Glucosidases
Hydration
Candida tropicalis
Arthrobacter
Substrates
Enzymes
Catalysis
Hydroxyl Radical
Protons
Catalytic Domain
Sugars
Ions
Nuclear magnetic resonance
Candida
Demonstrations
Proton Magnetic Resonance Spectroscopy
carbohydrase
2,6-anhydro-1-deoxygluco-hept-1-enitol

ASJC Scopus subject areas

  • Biochemistry

Cite this

Scope and mechanism of carbohydrase action. Stereospecific hydration of 2,6-anhydro-1-deoxy-D-gluco-hept-1-enitol catalyzed by α- and β-glucosidases and an inverting exo-α-glucanase. / Hehre, Edward J.; Brewer, Curtis F.; Uchiyama, Takao; Schlesselmann, Peter; Lehmann, Jochen.

In: Biochemistry, Vol. 19, No. 15, 1980, p. 3557-3564.

Research output: Contribution to journalArticle

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T1 - Scope and mechanism of carbohydrase action. Stereospecific hydration of 2,6-anhydro-1-deoxy-D-gluco-hept-1-enitol catalyzed by α- and β-glucosidases and an inverting exo-α-glucanase

AU - Hehre, Edward J.

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AB - D-Glucosylases of several types have been found to catalyze specific hydration and glycosyl transfer reactions (the former predominating) when incubated with 2,6-anhydro-1-deoxy-D-gluco-hept-1-enitol. α-Glucosidases from Candida tropicalis and from rice convert this substrate into 1-deoxy-α-D-gluco-heptulose. The β-glucosidase from sweet almonds and an exo-1,6-glucanase (glucodextranase) from Arthrobacter globiformis convert the substrate to 1-deoxy-β-D-gluco-heptulose. The anomeric form of the sugar was identified by following 1H NMR spectra of enzyme-enitol digests. The C-1 methyl group protons of the α and β anomers resonated at 1.45 and 1.40 ppm, respectively. Nonenzymatic anomerization of the β to the α form of the sugar was noted and, at equilibrium, 13C and hydroxyl 1H NMR spectra showed the presence of only the α anomer. The present work provides the first demonstration of the ability of an "inverting" glucanase to create specific anomeric configuration de novo from a substrate lacking α or β configuration. Indeed, the actions of glucodextranase on this enolic glycosyl donor lead not only to 1-deoxy-β-D-gluco-heptulose but also to a 1-deoxy-α-D-gluco-heptulosyl transfer product when enitol is the acceptor. These results reveal a catalytic mechanism that is able to convert a substrate to products of either configuration, depending on the type of acceptor. A mechanism is proposed for the stereospecific hydration of 2,6-anhydro-1-deoxy-D-gluco-hept-1-enitol by each enzyme, involving an incipient glycosyl carbonium ion and assuming the presence at the active site of two carboxyl groups arranged to account for catalysis of reactions with appropriate glycosidic substrates. These observations provide strong evidence for the functional flexibility of the catalytic groups of glycosylases.

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