Synthetic Variable Domain Glycopeptides for Neutralizing Epitope Characterization

描述（由申请人提供）：HIV-1包膜上广泛中和表位的鉴定和表征是设计有效HIV-1疫苗的重要步骤。最近，一类新的广泛中和抗体（bNAbs），包括PG9、PG16和PGT类抗体，已经从hiv感染的“精英控制者”中分离出来。这些bnab以显著的广度和效力中和原代HIV-1毒株。这些bnab抗原识别的一个共同特征是它们都靶向gp120 V1/V2和/或V3区域的聚糖依赖的四级表位。最近的x射线晶体结构研究表明，PG9在V1/V2结构域的N160和N156位点与n -聚糖结合，PGT128在V3结构域的N322和N301位点识别保守的n -聚糖。然而，中和表位的确切性质，特别是N156和N301上n -聚糖的精细结构仍有待表征。由于HIV-1 gp120糖基化的复杂性和异质性，进一步的表位定位变得复杂。我们假设独特的V1/V2和V3糖肽构成了这些bnab的中和表位。为了验证这一假设，我们将进行两个具体目标中描述的实验。目的1是通过一种新的化学酶方法，设计和合成具有N-糖基化位点上固定N-聚糖的环状V1/V2和V3 HIV-1糖肽。目的2是通过与合成的糖肽的结合和结构研究来表征中和抗体对抗原的识别。此外，合成的糖肽将用于检测hiv感染“非进展者”血清中糖依赖性、V1/V2和v3特异性中和抗体。这些研究可能为HIV-1疫苗设计提供重要的见解。

DESCRIPTION (provided by applicant): Identification and characterization of broadly neutralizing epitopes on the HIV-1 envelope are important steps in the design of an effective HIV-1 vaccine. Recently, a new class of broadly neutralizing antibodies (bNAbs), including PG9, PG16, and the PGT class antibodies, has been isolated from HIV-infected "elite controllers". These bNAbs neutralize primary HIV-1 strains with remarkable breadth and potency. A common feature of antigen recognition by these bNAbs is that they all target glycan-dependent quaternary epitopes at the V1/V2 and/or V3 regions of gp120. Recent X-ray crystal structural studies indicate that PG9 binds to N-glycans at N160 and N156 in the context of V1/V2 domain, and PGT128 recognizes conserved N-glycans at N322 and N301 sites in the context of V3 domain. However, the precise nature of the neutralizing epitopes, particularly the fine structures of the N-glycans at N156 and N301 remains to be characterized. Further mapping of the epitopes is complicated by the complexity and heterogeneity of glycosylation of HIV-1 gp120. We hypothesize that unique V1/V2 and V3 glycopeptides constitute the neutralizing epitopes for these bNAbs. To test this hypothesis, we will perform experiments described in two specific aims. Aim 1 is to design and synthesize cyclic V1/V2 and V3 HIV-1 glycopeptides with defined N-glycans being attached at the conserved N- glycosylation sites, by a novel chemoenzymatic method. Aim 2 is to characterize antigen recognition by the neutralizing antibodies through binding and structural studies with the synthetic glycopeptides. In addition, the synthetic glycopeptides will be used to detect glycan- dependent, V1/V2 and V3-specific neutralizing antibodies in sera from HIV-infected "non- progressors". These studies are likely to provide important insights for HIV-1 vaccine design. PUBLIC HEALTH RELEVANCE: HIV/AIDS is a serious global epidemic that threatens human health and social stability. The proposed research aims to characterize the neutralizing epitopes of broadly neutralizing antibodies, which will provide important insights in HIV-1 vaccine design.