Toneguzzo F, Ghosh H P
Proc Natl Acad Sci U S A. 1978 Feb;75(2):715-9. doi: 10.1073/pnas.75.2.715.
Translation in vitro of the mRNA coding for the vesicular stomatitis virus membrane glycoprotein G in a membrane-free ribosomal extract from HeLa cells allowed the synthesis of only the unglycosylated protein G1 (molecular weight, 63,000). Addition of stripped crude microsomal membranes from HeLa cells resulted in the conversion of G1 to the glycosylated protein G2 (molecular weight, 67,000). The G2 protein synthesized by the reconstructed microsomal membrane/ribosome system was found to be segregated inside the microsomal membrane vesicles and was thus protected from the proteolytic action of trypsin and chymotrypsin. Stripped membranes were required at an early stage of protein synthesis for the synthesized protein to be inserted into the membrane vesicles and to be glycosilated. The segregated protein G2, however, was not completely protected from proteolytic digestion, showing that a portion of the polypeptide chain of about 3000 daltons was present on the cytoplasmic side of the membrane vesicle. Our data thus suggest that, unlike the secretory proteins, the membrane glycoproteins are not completely discharged across the microsomal membranes.
在来自HeLa细胞的无膜核糖体提取物中对编码水泡性口炎病毒膜糖蛋白G的mRNA进行体外翻译，结果仅合成了未糖基化的蛋白G1（分子量为63,000）。添加来自HeLa细胞的脱垢粗微粒体膜导致G1转化为糖基化蛋白G2（分子量为67,000）。发现由重建的微粒体膜/核糖体系统合成的G2蛋白被分隔在微粒体膜泡内部，因此免受胰蛋白酶和胰凝乳蛋白酶的蛋白水解作用。在蛋白质合成的早期阶段需要脱垢膜，以便将合成的蛋白质插入膜泡并进行糖基化。然而，分隔的蛋白G2并未完全免受蛋白水解消化，这表明约3000道尔顿的多肽链的一部分存在于膜泡的细胞质侧。因此，我们的数据表明，与分泌蛋白不同，膜糖蛋白不会完全穿过微粒体膜排出。
