Fennie C, Lasky L A
Department of Molecular Immunology, Genentech, Inc., South San Francisco, California 94080.
J Virol. 1989 Feb;63(2):639-46. doi: 10.1128/JVI.63.2.639-646.1989.
The intracellular folding of the human immunodeficiency virus type 1 gp120 has been assessed by analyzing the ability of the glycoprotein to bind to the viral receptor CD4. Pulse-chase experiments revealed that the glycoprotein was initially produced in a conformation that was unable to bind to CD4 and that the protein attained the appropriate tertiary structure for binding with a half-life of approximately 30 min. The protein appears to fold within the rough endoplasmic reticulum, since blocking of transport to the Golgi apparatus by the oxidative phosphorylation inhibitor carbonyl cyanide m-chlorophenylhydrazone did not appear to perturb the folding kinetics of the molecule. The relatively lengthy folding time was not due to modification of the large number of N-linked glycosylation sites on gp120, since inhibition of the first steps in oligosaccharide modification by the inhibitors deoxynojirimycin or deoxymannojirimycin did not impair the CD4-binding activity of the glycoprotein. However, production of the glycoprotein in the presence of tunicamycin and removal of the N-linked sugars by endoglycosidase H treatment both resulted in deglycosylated proteins that were unable to bind to CD4, suggesting in agreement with previous results, that glycosylation contributes to the ability of gp120 to bind to CD4. Interestingly, incomplete endoglycosidase H treatment revealed that a partially glycosylated glycoprotein could bind to the receptor, implying that a subset of glycosylation sites, perhaps some of those conserved in different isolates of human immunodeficiency virus type 1, might be important for binding of the viral glycoprotein to the CD4 receptor.
通过分析糖蛋白与病毒受体CD4结合的能力，对人类免疫缺陷病毒1型gp120的细胞内折叠进行了评估。脉冲追踪实验表明，该糖蛋白最初以无法与CD4结合的构象产生，并且该蛋白以约30分钟的半衰期获得了与CD4结合的合适三级结构。该蛋白似乎在粗面内质网中折叠，因为氧化磷酸化抑制剂羰基氰化物间氯苯腙阻断向高尔基体的转运似乎并未干扰该分子的折叠动力学。相对较长的折叠时间并非由于gp120上大量N-连接糖基化位点的修饰，因为脱氧野尻霉素或脱氧甘露野尻霉素抑制寡糖修饰的第一步并未损害该糖蛋白的CD4结合活性。然而，在衣霉素存在下产生糖蛋白并通过内切糖苷酶H处理去除N-连接糖，均导致去糖基化蛋白无法与CD4结合，这与先前的结果一致，表明糖基化有助于gp120与CD4结合的能力。有趣的是，不完全的内切糖苷酶H处理表明，部分糖基化的糖蛋白可以与受体结合，这意味着一部分糖基化位点，可能是人类免疫缺陷病毒1型不同分离株中一些保守的位点，可能对病毒糖蛋白与CD4受体的结合很重要。
