Crystal Structures of Two Immune Complexes Identify Determinants for Viral Infectivity and Type-Specific Neutralization of Human Papillomavirus.
Crystal Structures of Two Immune Complexes Identify Determinants for Viral Infectivity and Type-Specific Neutralization of Human Papillomavirus.
复制标题
两种免疫复合物的晶体结构确定了病毒感染性和人乳头瘤病毒类型特异性中和的决定因素
作者：
Li Z;Wang D;Gu Y;Song S;He M;Shi J;Liu X;Wei S;Li J;Yu H;Zheng Q;Yan X;Baker TS;Zhang J;McLellan JS;Li S;Xia N
摘要持续的高危型人乳头瘤病毒（HPV）感染是宫颈癌的主要病因。由仅L1的病毒样颗粒（VLP）引起的中和抗体可以阻断HPV感染;然而，缺乏高分辨率结构限制了我们对病毒感染模式的理解以及在分子水平上对类型特异性的要求。在此，我们描述了两种抗体A12 A3和28 F10，其通过两种不同的结合化学计量分别特异性结合并中和HPV 58和HPV 59。我们发现A12 A3的表位聚集在两个相邻的HPV 58 L1单体的DE环中，而28 F10识别单个单体的HPV 59 FG环。通过基于结构的诱变和抗体结合分析，我们进一步鉴定了HPV 58 D154、S168和N170以及HPV 59 M267、Q270、E273、Y276、K278和R283残基，这些残基在病毒感染中起关键作用。通过将这些战略性表位残基替换到其他HPV基因型中，我们可以将抗体的类型特异性结合重定向到这些基因型，从而突出这些特异性残基HPV 58 R161、S168和N308以及HPV 59 Q270、E273和D281的重要性。总的来说，我们的研究结果提供了对感染性和类型特异性所需的HPV潜在结构决定因素的分子见解。高危型人乳头瘤病毒（HPV）被认为是影响上皮粘膜的癌症（如宫颈癌）的主要致病病原体。然而，由于缺乏中和位点的高分辨率结构信息，我们尚未确定HPV感染的精确模式以及不同类型的HPV如何引起感染。我们在这项研究中的晶体结构揭示了两种不同抗体的离散结合化学计量。我们显示一个A12 A3 Fab结合到一个HPV 58五聚体的中心，而五个28 F10 Fab沿着一个HPV 59五聚体的顶部边缘结合。此外，通过有针对性的表位分析，我们表明，6至7个不连续的残基的L1主要衣壳蛋白的HPV的决定因素，至少部分，病毒感染和类型特异性。这些知识将帮助我们解开HPV感染的过程，并可能用于推动靶向中和敏感部位的治疗方法的开发。高危型人乳头瘤病毒（HPV）被认为是影响上皮粘膜的癌症（如宫颈癌）的主要致病病原体。然而，由于缺乏中和位点的高分辨率结构信息，我们尚未确定HPV感染的精确模式以及不同类型的HPV如何引起感染。我们在这项研究中的晶体结构揭示了两种不同抗体的离散结合化学计量。我们显示一个A12 A3 Fab结合到一个HPV 58五聚体的中心，而五个28 F10 Fab沿着一个HPV 59五聚体的顶部边缘结合。此外，通过有针对性的表位分析，我们表明，6至7个不连续的残基的L1主要衣壳蛋白的HPV的决定因素，至少部分，病毒感染和类型特异性。这些知识将帮助我们解开HPV感染的过程，并可能用于推动靶向中和敏感部位的治疗方法的开发。
ABSTRACT Persistent, high-risk human papillomavirus (HPV) infection is the primary cause of cervical cancer. Neutralizing antibodies elicited by L1-only virus-like particles (VLPs) can block HPV infection; however, the lack of high-resolution structures has limited our understanding of the mode of virus infection and the requirement for type specificity at the molecular level. Here, we describe two antibodies, A12A3 and 28F10, that specifically bind to and neutralize HPV58 and HPV59, respectively, through two distinct binding stoichiometries. We show that the epitopes of A12A3 are clustered in the DE loops of two adjacent HPV58 L1 monomers, whereas 28F10 recognizes the HPV59 FG loop of a single monomer. Via structure-based mutagenesis and analysis of antibody binding, we further identified the residues HPV58 D154, S168, and N170 and HPV59 M267, Q270, E273, Y276, K278, and R283, which play critical roles in virus infection. By substituting these strategic epitope residues into other HPV genotypes, we could then redirect the type-specific binding of the antibodies to these genotypes, thus highlighting the importance of these specific residues, HPV58 R161, S168, and N308 and HPV59 Q270, E273, and D281. Overall, our findings provide molecular insights into potential structural determinants of HPV required for infectivity and type specificity. IMPORTANCE High-risk human papillomaviruses (HPVs) are considered the major causative pathogens of cancers that affect epithelial mucosa, such as cervical cancer. However, because of the lack of high-resolution structural information on the sites of neutralization, we have yet to determine the precise mode of HPV infection and how different types of HPV cause infection. Our crystal structures in this study have uncovered discrete binding stoichiometries for two different antibodies. We show that one A12A3 Fab binds to the center of one HPV58 pentamer, whereas five 28F10 Fabs bind along the top fringe of one HPV59 pentamer. Furthermore, through targeted epitope analysis, we show that 6 to 7 discontinuous residues of the L1 major capsid protein of HPV are determinants, at least in part, for virus infection and type specificity. This knowledge will help us to unravel the process of HPV infection and can potentially be used to drive the development of therapeutics that target neutralization-sensitive sites. IMPORTANCE High-risk human papillomaviruses (HPVs) are considered the major causative pathogens of cancers that affect epithelial mucosa, such as cervical cancer. However, because of the lack of high-resolution structural information on the sites of neutralization, we have yet to determine the precise mode of HPV infection and how different types of HPV cause infection. Our crystal structures in this study have uncovered discrete binding stoichiometries for two different antibodies. We show that one A12A3 Fab binds to the center of one HPV58 pentamer, whereas five 28F10 Fabs bind along the top fringe of one HPV59 pentamer. Furthermore, through targeted epitope analysis, we show that 6 to 7 discontinuous residues of the L1 major capsid protein of HPV are determinants, at least in part, for virus infection and type specificity. This knowledge will help us to unravel the process of HPV infection and can potentially be used to drive the development of therapeutics that target neutralization-sensitive sites.