Ballou L, Gopal P, Krummel B, Tammi M, Ballou C E
Proc Natl Acad Sci U S A. 1986 May;83(10):3081-5. doi: 10.1073/pnas.83.10.3081.
A mutant of Saccharomyces cerevisiae with the genotype mnn1 mnn2 mnn9 gls1 synthesizes mannoproteins with oligosaccharides having the composition Glc3Man10Glc-NAc2- owing to the mnn9 defect, which prevents synthesis of the outer chain, the mnn1 defect, which prevents branching of the core, and the gls1 mutation, which prevents deglucosylation of the resultant glycoprotein as a consequence of a defective glucosidase-I [Tsai, P.-K., Ballou, L., Esmon, B., Schekman, R. & Ballou, C. E. (1984) Proc. Natl. Acad. Sci. USA 81, 6340-6343]. (The mnn2 defect is not expressed in presence of the mnn9 mutation.) This strain spontaneously forms new colonies in which gls1 is suppressed owing to a defect in synthesis of dolichol phosphoglucose, the glucosylation substrate. The new mutant, designated mnn1 mnn2 mnn9 gls1 dpg1, synthesizes and secretes invertase (EC 3.2.1.26) that has a higher mobility on native gel electrophoresis than that made by the parent strain, the consequence of a reduction in both the size and the number of carbohydrate chains. The mannoprotein chains have the mnn1 mnn9 structure (Man10Glc-NAc2-), and the invertase is resolved by gel electrophoresis in sodium dodecyl sulfate into two major and two minor bands that represent homologs with about 4-7 carbohydrate units, in contrast to about 8-11 chains in the parent strain. Thus, the inability to glucosylate the lipid-linked precursor reduces the efficiency of glycosylation of the protein chains. The genetic defect is in synthesis of the glucose donor dolichol phosphoglucose, but the mutation is nonallelic with the reported alg5-1 mutation, which has a similar phenotype [Runge, K. W., Huffaker, T. C. & Robbins, P. W. (1984) J. Biol. Chem. 259, 412-417].
基因型为mnn1 mnn2 mnn9 gls1的酿酒酵母突变体合成的甘露糖蛋白带有组成成分为Glc3Man10Glc-NAc2-的寡糖，这是由于mnn9缺陷（阻止外链合成）、mnn1缺陷（阻止核心分支）以及gls1突变（由于葡糖苷酶I缺陷导致合成的糖蛋白无法进行去糖基化）所致[蔡，P.-K.，巴卢，L.，埃斯蒙，B.，谢克曼，R. & 巴卢，C. E.（1984年）《美国国家科学院院刊》81，6340 - 6343]。（在mnn9突变存在时，mnn2缺陷不表现。）该菌株会自发形成新菌落，其中gls1因多萜醇磷酸葡萄糖（糖基化底物）合成缺陷而受到抑制。新的突变体，命名为mnn1 mnn2 mnn9 gls1 dpg1，合成并分泌的转化酶（EC 3.2.1.26）在天然凝胶电泳上的迁移率高于亲本菌株所产生的转化酶，这是碳水化合物链的大小和数量减少的结果。甘露糖蛋白链具有mnn1 mnn9结构（Man10Glc-NAc2-），并且该转化酶在十二烷基硫酸钠凝胶电泳中可分离为两条主要带和两条次要带，这些带代表具有约4 - 7个碳水化合物单元的同系物，相比之下亲本菌株中有约8 - 11条链。因此，无法对脂质连接前体进行糖基化会降低蛋白质链糖基化的效率。遗传缺陷在于葡萄糖供体多萜醇磷酸葡萄糖的合成，但该突变与已报道的具有相似表型的alg5-1突变是非等位的[朗格，K. W.，哈法克，T. C. & 罗宾斯，P. W.（1984年）《生物化学杂志》259，412 - 417]。
