COMPARTMENTALIZED ASSEMBLY OF OLIGOSACCHARIDES ON EXPORTED GLYCOPROTEINS IN YEAST
COMPARTMENTALIZED ASSEMBLY OF OLIGOSACCHARIDES ON EXPORTED GLYCOPROTEINS IN YEAST
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DOI：
10.1016/0092-8674(81)90063-5
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发表时间：
1981-01-01
期刊：
影响因子：
64.5
通讯作者：
SCHEKMAN, R
中科院分区：
文献类型：
--
作者：
ESMON, B;NOVICK, P;SCHEKMAN, R
利用温度敏感的酿酒酵母分泌突变体(SEC)对糖蛋白低聚糖合成的阶段和定位进行了评价。在不允许的生长温度下(37℃。C)，sec突变体积累了分泌细胞器和糖蛋白。组织化学染色和切片EM显示，分泌的糖蛋白酸性磷酸酶存在于3个不同的细胞器中的一个中：内质网、被称为Berkeley小体的高尔基体结构和80-100 nm的囊泡。在37度的温度下生产。C、转化酶和酸性磷酸酶在积累内质网的突变株中的碳水化合物比其他突变株少，也比野生型菌株少。外源转化酶在十二烷基硫酸钠-聚丙烯酰胺凝胶上以异相形式迁移，表观相对分子质量为100-140kd[千道尔顿]。放射性标记蔗糖酶是从积累内质网突变细胞的提取物中免疫沉淀出来的，它以一组3种不同的蛋白质形式迁移，表观分子量分别为79、81和83kd；其他突变株产生的形式更像分泌酶。在每种情况下，用内切糖苷酶H处理去除N-糖基连接的低聚糖产生一个离散的物种，它以61kd的蛋白质的形式迁移。对突变体中积累的大量糖蛋白的免疫化学分析表明，N-连接的寡糖的主要部分，即外链，是在物质从内质网通过后添加的。
Temperature-sensitive secretory mutants (sec) of S. cerevisiae were used to evaluate the stages and localization of glycoprotein oligosaccharide synthesis. At the nonpermissive growth temperature (37.degree. C), the sec mutants accumulated secretory organelles and glycoproteins. Histochemical staining and thin-section EM revealed that the secreted glycoprotein, acid phosphatase, is contained within 1 of 3 distinct organelles that accumulate in different mutants: endoplasmic reticulum; Golgi-like structures called Berkeley bodies; and 80-100 nm vesicles. When produced at 37.degree. C, invertase and acid phosphatase have less carbohydrate in the mutants that accumulate ER than in other mutants, or than in the wild-type strain. External invertase migrates on SDS[sodium dodecyl sulfate]-polyacrylamide gels as a heterogeneous species with an apparent MW of 100-140 kd [kilodaltons]. Radiolabeled invertase, immunoprecipitated from extracts of ER-accumulating mutant cells, migrates as a set of 3 discrete protein species with apparent of 79, 81 and 83 kd; the other mutants produce a form more like the secreted enzyme. In each case, removal of N-glycosidically linked oligosaccharides by treatment with endoglycosidase H produces a discrete species that migrates as a protein of 61 kd. Immunochemical analysis of bulk glycoprotein accumulated in the mutants suggests that a major portion of the N-linked oligosaccharide, the outer chain, is added after material passes from the ER.