1996 Fiscal Year Annual Research Report

糸状菌結晶α-グリコシダーゼの基質認識機構及び糖鎖構造の解析

Research Project

Project/Area Number	06453164
Research Institution	HOKKAIDO UNIVERSITY
Principal Investigator	千葉誠哉北海道大学, 農学部, 教授 (30001449)
Keywords	α-グルコシダーゼ / α-ガラクトフラノース残基 / 第二次動力学的アイソトープ効果 / D-グルカ-ル水和反応 / 高マンノース型糖鎖
Research Abstract	A. niger結晶α-グルコシダーゼ(いわゆるマルターゼ型酵素)は二つのヘテロサブユニット(P1、227アミノ酸残基;P2、719アミノ酸残基)から構成されていた。両者間にS-S結合は存在しない。全一次構造はエドマン法とゲノムDNAの配列解析から明らかにされた。その結果、本酵素は一本のポリペプチド鎖としてP1続いてP2の順に翻訳された後に、プロテアーゼによる限定分解をうけて二つのサブユニットに構築されることが判明した。P2のAsp(224)が本酵素の触媒基の一つであることが推定された。7種の高マンノース型糖鎖を単離し、構造決定がなされた。4種の糖鎖はα-ガラクトフラノース残基を含み、そのうち、3種は新規構造であった。0-型糖鎖も単離され、構造解析の結果、マンノースとα-マンノシル-(1→2)-マンノースが本酵素のSerあるいはThrに直接結合することが推察された。これらの結果を基に、A. niger α-グルコシダーゼと他起源α-グルコシダーゼの一次構造の比較がなされ、α-グルコシダーゼが一次構造上から二つのファミリーに分けられることを明らかにした。また、α-グルコシダーゼとグルコアミラーゼによるD-グルカ-ルの水和反応の解析、α-第二次動力学的アイソトープ効果の解析を基に、これらの酵素の基質認識の仕組みが考察され反応機構モデルが提案された。

Research Products
(25 results)

All Other

All Publications (25 results)

[Publications] T. Takayanagi: "Identification of Ionizable Groups Essential to the Activity of Isomalto-dextranase from Arthrobacter globiformis" Biosci. Biotech. Biochem.58(11). 1995-1999 (1994)
[Publications] H. Kawagishi: "A Sialic Acid-binding Lectin from the Mashroom Hericium erinaceum" FEBS Lett.340(1). 56-58 (1994)
[Publications] T. Takayanagi: "Novel Structures of N-linked High-mannose Type Oligosaccharides Containing α-D-Galactofuranosyl Linkages in Aspergillus niger α-D-Glucosidase" Carbohydr. Res.256(1). 149-158 (1994)
[Publications] A. Iwai: "Molecular Cloning and Expression of an Isomalto-dextranase Gene from Arthrobactor globiformis" J. Bacteriol.176(24). 7730-7734 (1994)
[Publications] H. Kawagishi: "Two Lectins from Marine Sponge Halichondria okadai" J. Biol. Chem.269(2). 1375-1379 (1994)
[Publications] S. Onodera: "Substrate Specificity and Subsite Affinities of Rabbit Liver Acid α-Glucosidase" J. Biochem.116(11). 7-11 (1994)
[Publications] 高柳勉: "イソマルトデキストラナーゼによる酸処理デキストランからのイソマルトースの新しい調製法" 応用糖質科学. 42(4). 381-385 (1995)
[Publications] H. Mori: "Substrate Specificity and Subsite Affinities of α-Amylase from Germinating Cotyledons of Phaseolus vulgaris L. ev Toramame" Oyo Toshitsu Kagaku (J. Appl. Glycosci.). 42(4). 387-394 (1995)
[Publications] S. Iwanami: "Chemical Modification and Amino Acid Sequence of Active Site in Sugar Beet α-Glucosidase" Biosci. Biotech. Biochem.59(3). 459-463 (1995)
[Publications] 千葉誠哉: "α-グルコシダーゼとグルコアミラーゼ" 農化. 69(8). 1050-1054 (1995)
[Publications] H. Kawagishi: "Two N-acetyl-D-galactosamine-specific Lectins, PAL-I and PAL-II, from the Mushroom Phaeolepiota aurea" Phytochemistry. 41(4). 1013-1016 (1996)
[Publications] S. Iwanami: "Identification of Essential Ionizable Groups of Rice α-Glucosidase II" Oyo Toshitsu Kagaku (J. Appl. Glycosci.). 43(1). 67-71 (1996)
[Publications] 千葉誠哉: "トレハロース生成酵素について" 応用糖質科学. 43(2). 223-226 (1996)
[Publications] T. Mizuno: "Reinvestigation of Isomltotrio-dextranase from Brevibacterium fuscum var. dextranlycum" Oyo Toshitsu Kagaku (J. Appl. Glycosci.). 43(3). 347-353 (1996)
[Publications] A. Kita: "Enzymatic Properties of Three α-Glucosidases from Aspergillus oryzae" Oyo Toshitsu Kagaku (J. Appl. Glycosci.). 43(3). 325-330 (1996)
[Publications] R. Surarit: "Chemical Modification of β-Glucosidase/β-Fucosidase from Dalbergia Cochinchinensis Picrre by Conduritol B Epoxide" Biosci. Biotech. Biochem.60(8). 1265-1268 (1996)
[Publications] S. Onodera: "Molecular Cloning and Nucleotide Sequences of cDNA and Gene Encoding endo-Inulinase from Penicillium purpurogenum" Biosci. Biotech. Biochem.60(11). 1780-1785 (1996)
[Publications] H. Kawagishi: "A Lectin from Mycelia of the Fungus, Ganoderma lucidum" Phytochemistry. 44(1). 7-10 (1997)
[Publications] R. Surarit: "Evidence for the Presence of a Single Active Site in β-D-Fucosidase/β-D-Glucosidase from Dalbergia Cochinchinensis Seeds" Biosci. Biotech. Biochem.61(1). 93-95 (1997)
[Publications] A. Kimura: "Identification of Essential Ionizable Groups in Active Site of Aspergillus niger α-Glucosidase" Biosci. Biotech. Biochem.61(3). 475-479 (1997)
[Publications] A. Nakamura: "Cloning and Sequencing of an α-Glucosidase Gene from Aspergillus niger and Its Expression in A. nidulans" J. Biotechnol.(in press). (1997)
[Publications] 木村淳夫: "Aspergillus niger 結晶α-グルコシダーゼの一次構造と糖鎖" 応用糖質科学. 44(2)(印刷中). (1997)
[Publications] 松井博和: "テンサイα-グルコシダーゼの基質特異性と全一次構造" 応用糖質科学. 44(2)(印刷中). (1997)
[Publications] S. Chiba: "Enzyme Chemistry and Molecular Biology of Amylases and Related Enzymes" CRC Press. Inc., 16 (1995)
[Publications] 千葉誠哉: "新しい酵素研究法" 東京化学同人, 13 (1995)

1996 Fiscal Year Annual Research Report

糸状菌結晶α-グリコシダーゼの基質認識機構及び糖鎖構造の解析

Principal Investigator

千葉 誠哉 北海道大学, 農学部, 教授 (30001449)

Research Products

[Publications] T. Takayanagi: "Identification of Ionizable Groups Essential to the Activity of Isomalto-dextranase from Arthrobacter globiformis" Biosci. Biotech. Biochem.58(11). 1995-1999 (1994)

[Publications] H. Kawagishi: "A Sialic Acid-binding Lectin from the Mashroom Hericium erinaceum" FEBS Lett.340(1). 56-58 (1994)

[Publications] T. Takayanagi: "Novel Structures of N-linked High-mannose Type Oligosaccharides Containing α-D-Galactofuranosyl Linkages in Aspergillus niger α-D-Glucosidase" Carbohydr. Res.256(1). 149-158 (1994)

[Publications] A. Iwai: "Molecular Cloning and Expression of an Isomalto-dextranase Gene from Arthrobactor globiformis" J. Bacteriol.176(24). 7730-7734 (1994)

[Publications] H. Kawagishi: "Two Lectins from Marine Sponge Halichondria okadai" J. Biol. Chem.269(2). 1375-1379 (1994)

[Publications] S. Onodera: "Substrate Specificity and Subsite Affinities of Rabbit Liver Acid α-Glucosidase" J. Biochem.116(11). 7-11 (1994)

[Publications] 高柳勉: "イソマルトデキストラナーゼによる酸処理デキストランからのイソマルトースの新しい調製法" 応用糖質科学. 42(4). 381-385 (1995)

[Publications] H. Mori: "Substrate Specificity and Subsite Affinities of α-Amylase from Germinating Cotyledons of Phaseolus vulgaris L. ev Toramame" Oyo Toshitsu Kagaku (J. Appl. Glycosci.). 42(4). 387-394 (1995)

[Publications] S. Iwanami: "Chemical Modification and Amino Acid Sequence of Active Site in Sugar Beet α-Glucosidase" Biosci. Biotech. Biochem.59(3). 459-463 (1995)

[Publications] 千葉誠哉: "α-グルコシダーゼとグルコアミラーゼ" 農化. 69(8). 1050-1054 (1995)

[Publications] H. Kawagishi: "Two N-acetyl-D-galactosamine-specific Lectins, PAL-I and PAL-II, from the Mushroom Phaeolepiota aurea" Phytochemistry. 41(4). 1013-1016 (1996)

[Publications] S. Iwanami: "Identification of Essential Ionizable Groups of Rice α-Glucosidase II" Oyo Toshitsu Kagaku (J. Appl. Glycosci.). 43(1). 67-71 (1996)

[Publications] 千葉誠哉: "トレハロース生成酵素について" 応用糖質科学. 43(2). 223-226 (1996)

[Publications] T. Mizuno: "Reinvestigation of Isomltotrio-dextranase from Brevibacterium fuscum var. dextranlycum" Oyo Toshitsu Kagaku (J. Appl. Glycosci.). 43(3). 347-353 (1996)

[Publications] A. Kita: "Enzymatic Properties of Three α-Glucosidases from Aspergillus oryzae" Oyo Toshitsu Kagaku (J. Appl. Glycosci.). 43(3). 325-330 (1996)

[Publications] R. Surarit: "Chemical Modification of β-Glucosidase/β-Fucosidase from Dalbergia Cochinchinensis Picrre by Conduritol B Epoxide" Biosci. Biotech. Biochem.60(8). 1265-1268 (1996)

[Publications] S. Onodera: "Molecular Cloning and Nucleotide Sequences of cDNA and Gene Encoding endo-Inulinase from Penicillium purpurogenum" Biosci. Biotech. Biochem.60(11). 1780-1785 (1996)

[Publications] H. Kawagishi: "A Lectin from Mycelia of the Fungus, Ganoderma lucidum" Phytochemistry. 44(1). 7-10 (1997)

[Publications] R. Surarit: "Evidence for the Presence of a Single Active Site in β-D-Fucosidase/β-D-Glucosidase from Dalbergia Cochinchinensis Seeds" Biosci. Biotech. Biochem.61(1). 93-95 (1997)

[Publications] A. Kimura: "Identification of Essential Ionizable Groups in Active Site of Aspergillus niger α-Glucosidase" Biosci. Biotech. Biochem.61(3). 475-479 (1997)

[Publications] A. Nakamura: "Cloning and Sequencing of an α-Glucosidase Gene from Aspergillus niger and Its Expression in A. nidulans" J. Biotechnol.(in press). (1997)

[Publications] 木村淳夫: "Aspergillus niger 結晶α-グルコシダーゼの一次構造と糖鎖" 応用糖質科学. 44(2)(印刷中). (1997)

[Publications] 松井博和: "テンサイα-グルコシダーゼの基質特異性と全一次構造" 応用糖質科学. 44(2)(印刷中). (1997)

[Publications] S. Chiba: "Enzyme Chemistry and Molecular Biology of Amylases and Related Enzymes" CRC Press. Inc., 16 (1995)

[Publications] 千葉誠哉: "新しい酵素研究法" 東京化学同人, 13 (1995)

千葉誠哉北海道大学, 農学部, 教授 (30001449)