Go Eden P, Chang Qing, Liao Hua-Xin, Sutherland Laura L, Alam S Munir, Haynes Barton F, Desaire Heather
Department of Chemistry, University of Kansas, Lawrence, KS 66045, USA.
J Proteome Res. 2009 Sep;8(9):4231-42. doi: 10.1021/pr9002728.
The extensive glycosylation of HIV-1 envelope proteins (Envs), gp120/gp41, is known to play an important role in evasion of host immune response by masking key neutralization epitopes and presenting the Env glycosylation as "self" to the host immune system. The Env glycosylation is mostly conserved but continues to evolve to modulate viral infectivity. Thus, profiling Env glycosylation and distinguishing interclade and intraclade glycosylation variations are necessary components in unraveling the effects of glycosylation on Env's immunogenicity. Here, we describe a mass spectrometry-based approach to characterize the glycosylation profiles of two rVV-expressed clade C Envs by identifying the glycan motifs on each glycosylation site and determining the degree of glycosylation site occupancy. One Env is a wild-type Env, while the other is a synthetic "consensus" Env (C.CON). The observed differences in the glycosylation profiles between the two clade C Envs show that C.CON has more unutilized sites and high levels of high mannose glycans; these features mimic the glycosylation profile of a Group M consensus immunogen, CON-S. Our results also reveal a clade-specific glycosylation pattern. Discerning interclade and intraclade glycosylation variations could provide valuable information in understanding the molecular differences among the different HIV-1 clades and in designing new Env-based immunogens.
已知HIV-1包膜蛋白（Env）gp120/gp41的广泛糖基化在逃避宿主免疫反应中起重要作用，它通过掩盖关键中和表位并将Env糖基化作为“自身”呈现给宿主免疫系统。Env糖基化大多是保守的，但仍在不断进化以调节病毒感染性。因此，分析Env糖基化并区分不同分支间和分支内的糖基化变异是阐明糖基化对Env免疫原性影响的必要组成部分。在此，我们描述了一种基于质谱的方法，通过识别每个糖基化位点上的聚糖基序并确定糖基化位点占据程度，来表征两种重组痘苗病毒（rVV）表达的C组Env的糖基化谱。一种Env是野生型Env，另一种是合成的“共识”Env（C.CON）。观察到的两种C组Env糖基化谱的差异表明，C.CON有更多未利用的位点和高水平的高甘露糖聚糖；这些特征模仿了M组共识免疫原CON-S的糖基化谱。我们的结果还揭示了一种分支特异性糖基化模式。识别不同分支间和分支内的糖基化变异可为理解不同HIV-1分支间的分子差异以及设计基于Env的新型免疫原提供有价值的信息。
