Shakin-Eshleman S H, Spitalnik S L, Kasturi L
Case Western Reserve University, School of Medicine, Cleveland, Ohio 44106, USA.
J Biol Chem. 1996 Mar 15;271(11):6363-6. doi: 10.1074/jbc.271.11.6363.
N-Linked glycosylation is a common form of protein processing that can profoundly affect protein expression, structure, and function. N-Linked glycosylation generally occurs at the sequon Asn-X-Ser/Thr, where X is any amino acid except Pro. To assess the impact of the X amino acid on core glycosylation, rabies virus glycoprotein variants were generated by site-directed mutagenesis with each of the 20 common amino acids substituted at the X position of an Asn-X-Ser sequon. The efficiency of core glycosylation at the sequon in each variant was quantified in a rabbit reticulocyte lysate cell-free translation system supplemented with canine pancreas microsomes. The presence of Pro at the X position completely blocked core glycosylation, whereas Trp, Asp, Chi, and Leu were associated with inefficient core glycosylation. The other variants were more efficiently glycosylated, and several were fully glycosylated. These findings demonstrate that the X amino acid is an important determinant of N-linked core-glycosylation efficiency.
N-连接糖基化是一种常见的蛋白质加工形式，可深刻影响蛋白质的表达、结构和功能。N-连接糖基化通常发生在序列Asn-X-Ser/Thr处，其中X是除脯氨酸以外的任何氨基酸。为了评估X氨基酸对核心糖基化的影响，通过定点诱变产生了狂犬病病毒糖蛋白变体，在Asn-X-Ser序列的X位置替换了20种常见氨基酸中的每一种。在补充有犬胰腺微粒体的兔网织红细胞裂解物无细胞翻译系统中，对每个变体中该序列处的核心糖基化效率进行了定量。X位置存在脯氨酸完全阻断了核心糖基化，而色氨酸、天冬氨酸、半胱氨酸和亮氨酸与低效的核心糖基化有关。其他变体的糖基化效率更高，有几种完全糖基化。这些发现表明，X氨基酸是N-连接核心糖基化效率的重要决定因素。
