Characterization of a specific alpha-mannosidase involved in oligosaccharide processing in Saccharomyces cerevisiae.
Characterization of a specific alpha-mannosidase involved in oligosaccharide processing in Saccharomyces cerevisiae.
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酿酒酵母中参与寡糖加工的特定 α-甘露糖苷酶的表征。
DOI：
10.1016/s0021-9258(18)89546-1
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发表时间：
1985
期刊：
影响因子：
--
通讯作者：
A. Herscovics
中科院分区：
文献类型：
--
作者：
S. Jelinek;T. Akiyama;B. Saunier;J. Tkacz;A. Herscovics
在0.5%Triton X-100存在下，在Sepharose 6 B上分级分离来自酿酒酵母X-2180的粗提取物，分离出两种含有α-甘露糖苷酶活性的酶级分。从凝胶中排除的级分I含有对作为底物的对硝基苯基-α-D-吡喃甘露糖苷和Man 9 GlcNAc寡糖的α-甘露糖苷酶活性，而包含在凝胶中的级分II仅含有寡糖α-甘露糖苷酶活性。后一种酶是非常特异的，并且从Man 9 GlcNAc中除去单个甘露糖残基，而级分I的α-甘露糖苷酶活性从Man 9 GlcNAc寡糖中除去几个甘露糖残基。在级分II的α-甘露糖苷酶存在下由Man 9 GlcNAc形成的Man 8 GlcNAc的高分辨率1H NMR分析仅显示具有以下结构的单一异构体：（见式;见正文）这种特异性酶最可能参与甘露糖蛋白生物合成期间寡糖的加工。1-脱氧野尻霉素的甘露糖类似物（50-500 μ M），双脱氧-1，5-亚氨基-D-甘露醇，抑制级分I和II的寡糖α-甘露糖苷酶活性的程度大致相同，但对作用于对硝基苯基-α-D-吡喃甘露糖苷的非特异性α-甘露糖苷酶没有影响。
Fractionation of a crude extract from Saccharomyces cerevisiae X-2180 on Sepharose 6B in the presence of 0.5% Triton X-100 resolves two enzyme fractions containing alpha-mannosidase activity. Fraction I which is excluded from the gel contains alpha-mannosidase activity toward both p-nitrophenyl-alpha-D-mannopyranoside and Man9GlcNAc oligosaccharide as substrates, whereas Fraction II which is included in the gel contains only oligosaccharide alpha-mannosidase activity. The latter enzyme is very specific and removes a single mannose residue from Man9GlcNAc, whereas the alpha-mannosidase activity of Fraction I removes several mannose residues from Man9GlcNAc oligosaccharide. High resolution 1H NMR analysis of the Man8GlcNAc formed from Man9GlcNAc in the presence of the alpha-mannosidase of Fraction II showed only a single isomer with the following structure: (see formula; see text) This specific enzyme is most probably involved in processing of oligosaccharide during biosynthesis of mannoproteins. The mannose analog of 1-deoxynojirimycin (50-500 microM), dideoxy-1,5-imino-D-mannitol, inhibits the oligosaccharide alpha-mannosidase activities of Fractions I and II to about the same extent, but has no effect on the nonspecific alpha-mannosidase which acts on p-nitrophenyl-alpha-D-mannopyranoside.