Structure-based approaches to antigen design utilize insights from antibody (Ab):antigen interactions and a refined understanding of protective Ab responses to engineer novel antigens presenting epitopes with conformations relevant to eliciting or discovering protective humoral responses. For human immunodeficiency virus-1 (HIV-1), one model of protection is provided by broadly neutralizing Abs (bnAbs) against epitopes present in the closed prefusion trimeric conformation of HIV-1 envelope glycoprotein, such as the variable loops 1-2 (V1V2) apex. Here, computational design and directed evolution yielded a novel V1V2 sequence variant with potential utility for inclusion in an immunogen for eliciting bnAbs, or as an epitope probe for their detection. The computational design goal was to engineer a minimal single-chain antigen with three copies of the V1V2 loops to support maintenance of closed prefusion V1V2 trimeric conformation and presentation of bnAb epitopes. Via directed evolution of this computationally designed single-chain antigen, we isolated a V1V2 sequence variant that in monomeric form exhibited preferential recognition by quaternary-preferring and conformation-dependent mAbs. Structural context and transferability of this phenotype to V1V2 sequences from all strains of HIV-1 tested suggest a conformation-stabilizing effect. This example demonstrates the potential utility of computational design and directed evolution-based protein engineering strategies to develop minimal, conformation-stabilized epitope-specific antigens.

中文翻译：

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迈向四元 HIV-1 表位的构象保真度：最小 V1V2 抗原的计算设计和定向进化。

基于结构的抗原设计方法利用抗体 (Ab):抗原相互作用的见解以及对保护性 Ab 反应的深入理解来设计新的抗原，该抗原呈递具有与引发或发现保护性体液反应相关的构象的表位。对于人类免疫缺陷病毒 1 (HIV-1)，一种保护模型是通过广泛中和 Abs (bnAbs) 来提供针对 HIV-1 包膜糖蛋白闭合预融合三聚体构象中存在的表位，例如可变环 1-2 ( V1V2) 顶点。在这里，计算设计和定向进化产生了一种新的 V1V2 序列变体，其具有潜在的用途，可包含在免疫原中以引发 bnAb，或作为其检测的表位探针。计算设计目标是设计具有三个 V1V2 环拷贝的最小单链抗原，以支持闭合前 V1V2 三聚体构象的维持和 bnAb 表位的呈递。通过这种计算设计的单链抗原的定向进化，我们分离出一种 V1V2 序列变体，该变体以单体形式表现出被四元偏好和构象依赖性单克隆抗体优先识别的能力。该表型的结构背景和从所有测试的 HIV-1 毒株的 V1V2 序列的可转移性表明具有构象稳定作用。这个例子展示了计算设计和基于定向进化的蛋白质工程策略在开发最小的、构象稳定的表位特异性抗原方面的潜在效用。