Preparation of High-Mannose type Oligosaccharides with mannosidases.

用甘露糖苷酶制备高甘露糖型寡糖。

• 批准号: 03660125
• 负责人: NAKAJIMA Tasuku
• 金额: $ 1.47万
• 依托单位: TOHOKU UNIVERSITY
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for General Scientific Research (C)
• 财政年份: 1990
• 资助国家: 日本
• 起止时间: 1990 至 1992
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-03660125/
• 关键词: Mannooligosaccharide; Mannosidase; alpha-1,2-mannosidase; alpha-1,3-mannanase; 高マンノース型オリゴ糖; α-1、3-マンノシダーゼ; α-1、3-マンノビオース; α-1、3-マンノオリゴ糖; α-1、3-マンナナーゼ; α-マンナナーゼ; αー1,2ーマンノシダ-ゼ; αー1,2ーマレノビオ-ス; 逆合成糖; αー1,6ーマンナナ-ゼ; αーマンノシダ-ゼ

The aim of this study is to develop the preparation method of the skeleton oligosaccharides of N-liked high-mannose type suger chains. First, we isolated a soil bacterium that produced alpha-1,2-mannosidase extracellulary. The enzyme was purified to homogeneity and characterized. The enzyme had a molecular weight of 380 Kda consisting of two homologous subunits of 190 Kda protein. The enzyme hydrolyzed specifically alpha-1,2-mannosidic linked oligosaccharides and D-mannose was liberated. The enzyme did not hydrolyze pNP- or methyl-alpha-mannoside. Using this mannosidase, the alpha-1,2-mannobiose and alpha-1,2-mannotriose were prepared by the reverce action of the enzyme with D-mannose as substrate. To obtainalpha-1,3-linked manno-oligosaccharides, we also found a new enzyme specific for alpha-1.3-mannan. A gram-negative bacterium, strain No. 9 was isolated from the soil samples and the organism secreted alpha-1,3-mannanase to produce alpha-1,3-manno-oligosaccharides. The enzyme was partially purified. The final product of the reaction was alpha-1,3-mannobiose. Above results indicate that alpha-1,2- and alpha-1,3-manno-oligosaccharides can be prepared enzymatically using the two bacterial enzymes.

本研究的目的是开发N型高甘露糖型糖链骨架寡糖的制备方法。首先，我们分离了一种产生胞外α-1,2-甘露糖苷酶的土壤细菌。将酶纯化至同质并进行表征。该酶的分子量为 380 Kda，由 190 Kda 蛋白质的两个同源亚基组成。该酶特异性水解α-1,2-甘露糖苷连接的寡糖并释放D-甘露糖。该酶不水解 pNP-或甲基-α-甘露糖苷。利用该甘露糖苷酶，以D-甘露糖为底物，通过酶的逆作用制备α-1,2-甘露二糖和α-1,2-甘露三糖。为了获得α-1,3-连接的甘露寡糖，我们还发现了一种针对α-1.3-甘露聚糖的新酶。从土壤样品中分离出革兰氏阴性细菌 9 号菌株，该生物体分泌 α-1,3-甘露聚糖酶以产生 α-1,3-甘露寡糖。该酶被部分纯化。反应的最终产物是α-1,3-甘露二糖。上述结果表明，可以使用两种细菌酶通过酶法制备α-1,2-和α-1,3-甘露寡糖。

项目成果

中島 佐(共同執筆): "糖鎖工学ーこれからの生命科学(植物・微生物の糖鎖工学)" 産業調査会事典出版センタ-, 32 (1992)

Satoshi Nakajima（合着）：《糖工程-未来生命科学（植物和微生物的糖工程）》工业研究会百科全书出版中心，32（1992）

中島 佑(分担執筆): "糖鎖工学" 産業調査会, 680 (1992)

Yu Nakajima（贡献者）：“糖工程”工业研究小组，680（1992）

丸山 穣,中島 佐,一島 英治: "αー1,2ーマンノシダ-ゼ生産菌の分離と酵素の性質について" 日本農芸化学会誌. 65. 293 (1991)

Jo Maruyama、Sa Nakajima、Eiji Kazushima：“α-1,2-甘露糖苷酶产生细菌的分离及其酶的特性”日本农业化学学会杂志 65. 293 (1991)。

Y.Chiba,Y.Yamagata T.Nakajima,E.Ichishima: "A New high-mannose type N-linbed Oligosaccharide from Aspergillus Carboxypeptidase." Biosci.Biotech.Biochem.56. 1371-1372 (1992)

Y.Chiba，Y.Yamagata T.Nakajima，E.Ichishima：“一种来自曲霉羧肽酶的新型高甘露糖型 N 线寡糖。”