Wibmer Constantinos Kurt, Gorman Jason, Ozorowski Gabriel, Bhiman Jinal N, Sheward Daniel J, Elliott Debra H, Rouelle Julie, Smira Ashley, Joyce M Gordon, Ndabambi Nonkululeko, Druz Aliaksandr, Asokan Mangai, Burton Dennis R, Connors Mark, Abdool Karim Salim S, Mascola John R, Robinson James E, Ward Andrew B, Williamson Carolyn, Kwong Peter D, Morris Lynn, Moore Penny L
Centre for HIV and STIs, National Institute for Communicable Diseases (NICD), of the National Health Laboratory Service (NHLS), Johannesburg, South Africa.
Faculty of Health Sciences, University of the Witwatersrand, Johannesburg, South Africa.
PLoS Pathog. 2017 Jan 11;13(1):e1006074. doi: 10.1371/journal.ppat.1006074. eCollection 2017 Jan.
A comprehensive understanding of the regions on HIV-1 envelope trimers targeted by broadly neutralizing antibodies may contribute to rational design of an HIV-1 vaccine. We previously identified a participant in the CAPRISA cohort, CAP248, who developed trimer-specific antibodies capable of neutralizing 60% of heterologous viruses at three years post-infection. Here, we report the isolation by B cell culture of monoclonal antibody CAP248-2B, which targets a novel membrane proximal epitope including elements of gp120 and gp41. Despite low maximum inhibition plateaus, often below 50% inhibitory concentrations, the breadth of CAP248-2B significantly correlated with donor plasma. Site-directed mutagenesis, X-ray crystallography, and negative-stain electron microscopy 3D reconstructions revealed how CAP248-2B recognizes a cleavage-dependent epitope that includes the gp120 C terminus. While this epitope is distinct, it overlapped in parts of gp41 with the epitopes of broadly neutralizing antibodies PGT151, VRC34, 35O22, 3BC315, and 10E8. CAP248-2B has a conformationally variable paratope with an unusually long 19 amino acid light chain third complementarity determining region. Two phenylalanines at the loop apex were predicted by docking and mutagenesis data to interact with the viral membrane. Neutralization by CAP248-2B is not dependent on any single glycan proximal to its epitope, and low neutralization plateaus could not be completely explained by N- or O-linked glycosylation pathway inhibitors, furin co-transfection, or pre-incubation with soluble CD4. Viral escape from CAP248-2B involved a cluster of rare mutations in the gp120-gp41 cleavage sites. Simultaneous introduction of these mutations into heterologous viruses abrogated neutralization by CAP248-2B, but enhanced neutralization sensitivity to 35O22, 4E10, and 10E8 by 10-100-fold. Altogether, this study expands the region of the HIV-1 gp120-gp41 quaternary interface that is a target for broadly neutralizing antibodies and identifies a set of mutations in the gp120 C terminus that exposes the membrane-proximal external region of gp41, with potential utility in HIV vaccine design.
全面了解广泛中和抗体靶向的HIV-1包膜三聚体区域，可能有助于合理设计HIV-1疫苗。我们之前在CAPRISA队列中鉴定出一名参与者CAP248，其在感染三年后产生了能够中和60%异源病毒的三聚体特异性抗体。在此，我们报告通过B细胞培养分离出单克隆抗体CAP248-2B，它靶向一个新的膜近端表位，包括gp120和gp41的元件。尽管最大抑制平台较低，通常低于50%抑制浓度，但CAP248-2B的广度与供体血浆显著相关。定点诱变、X射线晶体学和负染电子显微镜三维重建揭示了CAP248-2B如何识别一个依赖切割的表位，该表位包括gp120 C末端。虽然这个表位是独特的，但它在gp41的部分区域与广泛中和抗体PGT151、VRC34、35O22、3BC315和10E8的表位重叠。CAP248-2B具有构象可变的互补决定区，其轻链第三互补决定区异常长，有19个氨基酸。通过对接和诱变数据预测，环顶端的两个苯丙氨酸与病毒膜相互作用。CAP248-2B的中和作用不依赖于其表位附近的任何单个聚糖，低中和平台不能完全由N-或O-连接糖基化途径抑制剂、弗林蛋白酶共转染或与可溶性CD4预孵育来解释。病毒对CAP248-2B的逃逸涉及gp120-gp41切割位点的一组罕见突变。将这些突变同时引入异源病毒中可消除CAP248-2B的中和作用，但可使对35O22、4E10和10E8的中和敏感性提高10至100倍。总之，本研究扩展了HIV-1 gp120-gp41四级界面中作为广泛中和抗体靶点的区域，并鉴定出gp120 C末端的一组突变，这些突变暴露了gp41的膜近端外部区域，在HIV疫苗设计中具有潜在用途。
