Glycon specificity profiling of α-glucosidases using monodeoxy and mono-O-methyl derivatives of p-nitrophenyl α-d-glucopyranoside

Nishio, Takeshi; Hakamata, Wataru; Kimura, Atsuo; Chiba, Seiya; Takatsuki, Akira; Kawachi, Ryu; Oku, Tatsuo

doi:10.1016/s0008-6215(02)00026-5

Cited by 34 publications

(23 citation statements)

References 21 publications

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“…As shown in Fig. 2D, our combined reaction eliminates the plateau during glucose production with a Ͼ40-fold enhancement of hydration as com- (16). B, 31AG-catalyzed trans-addition to GlcAL (17,18).…”

Section: Resultsmentioning

confidence: 83%

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A Novel Metabolic Pathway for Glucose Production Mediated by α-Glucosidase-catalyzed Conversion of 1,5-Anhydrofructose

Kim

Saburi

Yu³

et al. 2012

Journal of Biological Chemistry

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Background: ␣-Glucosidase-catalyzed hydration on 1,5-anhydrofructose has not been clarified. Results: ␣-Glucosidases of glycoside hydrolase families 13 and 31 synthesize ␣-configurated product from 1,5-anhydrofructose. Combined reaction of glucan lyase and ␣-glucosidase enhances glucose production from starch. Conclusion: ␣-Glucosidases catalyze trans-addition on 1,5-anhydrofructose. Significance: Novel ␣-glucosidase-associated metabolic pathway of glucose formation from 1,5-anhydrofructose suggests a salvage process of unutilized 1,5-anhydrofructose to glucose in nature.

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Section: Resultsmentioning

confidence: 83%

“…31AGs display hydrolysis on p-nitrophenyl 2-deoxyglucoside ( Fig. 1A) (16) and hydration at a double bond of D-glucal (GlcAL) by trans-addition to produce ␣-2-deoxyglucose (␣2dGlc; Fig. 1B) (17,18), although 13AGs cannot catalyze those reactions (17).…”

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confidence: 99%

A Novel Metabolic Pathway for Glucose Production Mediated by α-Glucosidase-catalyzed Conversion of 1,5-Anhydrofructose

Kim

Saburi

Yu³

et al. 2012

Journal of Biological Chemistry

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“…Previously, we had elucidated the Km values of the enzyme for these three substrates at pH 4.0, and confirmed values of 0.59, 6.09 and 10.2 mM for Glc O pNP, 2D Glc O pNP and 3D Glc O pNP, respectively. 8) Taking into consideration these Km values and solubility of the deoxy substrates, pH dependency of the enzyme activity against these substrates was investigated at concentrations of 1 2 Km. Optimal pH for the ANGase reaction with Glc O pNP was 4.4, while the pH for both deoxy analogs was 5.6 (Fig.…”

Section: Resultsmentioning

confidence: 99%

“…9) Using substrates partially modified at their glycon portions, we have studied substrate specificities of various types of glycosidases. 5,6,8,14,17) Through these studies, we predicted that some properties of the hydrolytic actions of exo glycosidases are controlled by the glycon hydroxyl groups of their substrates. On the basis of this hypothesis, we investigated the role of the glycon hydroxyl groups of the glycoside during glycosidase action by kinetic studies on the hydrolysis of partially deoxygenated glycosides.…”

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confidence: 99%

Substrate Hydroxyl Groups Are Involved in the Ionization of Catalytic Carboxyl Groups of Aspergillus niger .ALPHA.-Glucosidase

真弘¹,

俊幸²,

航³

et al. 2004

J. Appl. Glycosci.

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Carbohydrate hydrolases are widely distributed in microorganisms, plants, insects, and mammals. These enzymes are classified on the basis of their modes of action (exo or endo type), sugar specificity, form of sugar chain, and configuration ( or anomer) and position of the glycosidic linkage. Hydrogen bonding interactions between the amino acids involved in the active site of these enzymes and the hydroxyl groups of their sugar substrates are important in the formation of the enzyme substrate (ES) complex. In fact, with some enzymes, systematic hydrogen bondings between their active sites and their substrates have been estimated from three dimen-s i o n a l structure analyses. For example, extensive hydrogen bond formation was indicated between the active site of glucoamylase (EC 3.2.1.3) from the mold Aspergillus niger and methyl maltoside or isomaltoside.1,2) Studies using mono deoxygenated substrates demonstrated that specific hydroxyl groups of the glycoside play an essential role in ES complex formation with glucoamylases, glucosidases ( EC 3.2.1.21), 10 13) galactosidases (EC 3.2.1.22), 6,14) galactosidases (EC 3.2.1.23), 15,16) and mannosidases (EC 3.2.1.24) 17) from various organisms. Active site amino acids involved in substrate binding have been identified by studies of site directed mutagenesis.18,19) Thus, investigations into interactions between an enzyme and its sugar substrate focus on hydrogen bond formation. However, Notenboom et al . reported that the nonreducing end sugar (glycon) OH 2 group of aryl cellobioside promotes transition state stabilization of reaction intermediates during hydrolytic reactions of the bacteria Cellulomonas fimi glucosidase. 20) Moreover, Frandesen et al . also indicated that OH 2 and 3 groups of the reducing end ring (aglycon) of the substrate methyl isomaltoside contribute to transition state stabilization in yeast glucosidase, while all glycon hydroxyl groups of the substrate are intimately involved in the formation of hydrogen bonds with the enzyme. 9) Using substrates partially modified at their glycon portions, we have studied substrate specificities of various types of glycosidases. 5,6,8,14,17) Through these studies, we predicted that some properties of the hydrolytic actions of exo glycosidases are controlled by the glycon hydroxyl groups of their substrates. On the basis of this hypothesis, we investigated the role of the glycon hydroxyl groups of the glycoside during glycosidase action by kinetic studies on the hydrolysis of partially deoxygenated glycosides.We chose A. niger glucosidase (ANGase) for this study because of its strong hydrolytic activity against 2 and 3 deoxy glucosides modified at the glycon portion, al-

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“…Kinetic studies of the hydrolysis of PNP α Glc, 1 and 2 were also carried out (Table 2). 9 Aglycon specificity profiling and inhibition of glucosidases using heptitol derivatives.…”

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N-Linked Oligosaccharide Processing Enzymes as Molecular Targets for Drug Discovery

Hakamata¹,

Muroi²,

Nishio³

et al. 2006

J. Appl. Glycosci.

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α Glucosidases (EC 3.2.1.20) are also exo acting carbohydrases, catalyzing the release of α D glucopyranose from the non reducing ends of various substrates, 1,2) and on the basis of amino acid sequence similarities, α glucosidases are classified into two families, family 13 and family 31. 3,4) Endoplasmic reticulum (ER) glucosidases, glucosidase I (EC 3.2.1.106) and glucosidase II (EC 3.2.1.84), are key enzymes in the biosynthesis of asparagine linked oligosaccharides that catalyze the first processing event after the transfer of Glc3Man9GlcNAc2 to proteins. These enzymes are a target for inhibition by anti viral agents that interfere with the formation of essential glycoproteins required in viral assembly, secretion and infectivity.5) Many papers reported that inhibitors of α glucosidases are potential therapeutics for the treatment of such diseases as viral diseases, cancer and diabetes. 5,6) However, many screenings of α glucosidase inhibitors did not use enzymes from target tissues or organs. We think that the molecular recognitions of three kinds of glucosidases (family 13, family 31 α glucosidases and ER glucosidases) are different. Therefore, the glycon and aglycon specificity profiling of glucosidases has been an important approach for the research of glucosidase inhibitors.In this research, we first describe the glycon and aglycon specificity profiling of glucosidases using small molecules as probes. Next, compounds designed and synthesized as glucosidase inhibitor candidates were evaluated with regard to their ability to inhibit three kinds of glucosidases. Finally, the glucosidase inhibitor candidates were tested for their anti viral activities in a cell culture system.Glycon specificity profiling of glucosidases using chemically modified substrates. Chemically modified substrates are effective methods in the study of substrate specificity profiling. We have applied this approach to family 13 and family 31 α glucosidases, 7 10) ER glucosidases, 11,12) α galactosidases 8,13) and α mannosidases 8,14) using partially substituted monosaccharides. We used all of the monodeoxy analogs of p nitrophenyl α D glucopyranoside (PNP α Glc) 1 4 (Fig. 1) as chemically modified substrates for glycon specificity profiling. We investigated the hydrolytic activities of family 13 and family 31 α glucosidases and ER glucosidase II of PNP α Glc and its deoxy derivatives 1 4, and checked the inhibitory activities of ER glucosidase I of PNP α Glc and probes 1 4, so that PNP α Glc was not a substrate for ER glucosidase I. These results are shown in Table 1. 11,12) Clearly, of the four deoxy derivatives of PNP α Glc 1 4, family 31 α glucosidases and ER glucosidase II hydrolyzed the 2 deoxy glucopyranoside (1); its activity with 1 appeared to be substantially higher than that with PNP α Glc. Kinetic studies of the hydrolysis of PNP α Glc, 1 and 2 were also carried out (Table 2) Abstract: N-Linked oligosaccharide processing enzymes are key enzymes in the biosynthesis of N-linked oligosaccharides. These enzymes are a molecular t...

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Glycon specificity profiling of α-glucosidases using monodeoxy and mono-O-methyl derivatives of p-nitrophenyl α-d-glucopyranoside

Cited by 34 publications

References 21 publications

A Novel Metabolic Pathway for Glucose Production Mediated by α-Glucosidase-catalyzed Conversion of 1,5-Anhydrofructose

A Novel Metabolic Pathway for Glucose Production Mediated by α-Glucosidase-catalyzed Conversion of 1,5-Anhydrofructose

Substrate Hydroxyl Groups Are Involved in the Ionization of Catalytic Carboxyl Groups of Aspergillus niger .ALPHA.-Glucosidase

N-Linked Oligosaccharide Processing Enzymes as Molecular Targets for Drug Discovery

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