Lüftenegger Daniel, Picard-Maureau Marcus, Stanke Nicole, Rethwilm Axel, Lindemann Dirk
Institut für Virologie, Medizinische Fakultät "Carl Gustav Carus," Technische Universität Dresden, Fetscherstr. 74, 01307 Dresden, Germany.
J Virol. 2005 Jun;79(12):7664-72. doi: 10.1128/JVI.79.12.7664-7672.2005.
The prototype foamy virus (PFV) glycoprotein, which is essential for PFV particle release, displays a highly unusual biosynthesis, resulting in posttranslational cleavage of the precursor protein into three particle-associated subunits, i.e., leader peptide (LP), surface (SU), and transmembrane (TM). Glycosidase digestion of metabolically labeled PFV particles revealed the presence of N-linked carbohydrates on all subunits. The differential sensitivity to specific glycosidases indicated that all oligosaccharides on LP and TM are of the high-mannose or hybrid type, whereas most of those attached to SU, which contribute to about 50% of its molecular weight, are of the complex type. Individual inactivation of all 15 potential N-glycosylation sites in PFV Env demonstrated that 14 are used, i.e., 1 out of 2 in LP, 10 in SU, and 3 in TM. Analysis of the individual altered glycoproteins revealed defects in intracellular processing, support of particle release, and infectivity for three mutants, having the evolutionarily conserved glycosylation sites N8 in SU or N13 and N15 in the cysteine-rich central "sheets-and-loops" region of TM inactivated. Examination of alternative mutants with mutations affecting glycosylation or surrounding sequences at these sites indicated that inhibition of glycosylation at N8 and N13 most likely is responsible for the observed replication defects, whereas for N15 surrounding sequences seem to contribute to a temperature-sensitive phenotype. Taken together these data demonstrate that PFV Env and in particular the SU subunit are heavily N glycosylated and suggest that although most carbohydrates are dispensable individually, some evolutionarily conserved sites are important for normal Env function of FV isolates from different species.
原型泡沫病毒（PFV）糖蛋白对PFV颗粒释放至关重要，其生物合成过程极为特殊，导致前体蛋白在翻译后裂解为三个与颗粒相关的亚基，即前导肽（LP）、表面蛋白（SU）和跨膜蛋白（TM）。对经代谢标记的PFV颗粒进行糖苷酶消化，结果显示所有亚基上均存在N-连接碳水化合物。对特定糖苷酶的不同敏感性表明，LP和TM上的所有寡糖均为高甘露糖型或杂合型，而附着在SU上的大多数寡糖（约占其分子量的50%）为复合型。对PFV Env中所有15个潜在N-糖基化位点进行逐个失活分析表明，其中14个位点被使用，即LP中的2个位点有1个被使用，SU中有10个，TM中有3个。对单个改变的糖蛋白进行分析发现，有三个突变体在细胞内加工、支持颗粒释放和感染性方面存在缺陷，这些突变体使SU中进化保守的糖基化位点N8或TM富含半胱氨酸的中央“片层-环”区域中的N13和N15失活。对影响这些位点糖基化或周围序列的替代突变体进行检测表明，N8和N13处糖基化的抑制最有可能是观察到的复制缺陷的原因，而对于N15，周围序列似乎导致了温度敏感表型。综合这些数据表明，PFV Env尤其是SU亚基高度N-糖基化，并且表明尽管大多数碳水化合物单个而言是可有可无的，但一些进化保守位点对于来自不同物种的FV分离株的正常Env功能很重要。
