Structural basis of HIV-1 neutralization by affinity matured Fabs directed against the internal trimeric coiled-coil of gp41.
Structural basis of HIV-1 neutralization by affinity matured Fabs directed against the internal trimeric coiled-coil of gp41.
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DOI：
10.1371/journal.ppat.1001182
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发表时间：
2010-11-11
期刊：
影响因子：
6.7
通讯作者：
Clore GM
中科院分区：
文献类型：
--
作者：
Gustchina E;Li M;Louis JM;Anderson DE;Lloyd J;Frisch C;Bewley CA;Gustchina A;Wlodawer A;Clore GM
HIV - 1 gp41的N - 七肽重复序列（N - HR）的保守内部三聚体卷曲螺旋在融合过程中通过形成发夹前体中间体而短暂暴露，因此是设计融合抑制剂和中和抗体的一个有吸引力的靶点。在先前的研究中，我们报道了一系列广泛中和的微型抗体，这些抗体是通过用人源组合抗体文库对三聚体N - HR卷曲螺旋模拟物进行淘选，然后利用CDR - H2环的靶向多样化进行亲和力成熟而从该文库中获得的。在此我们报道了N - HR模拟物5 - 螺旋与两种Fab片段的晶体结构，这两种Fab片段代表了该系列的两个极端：Fab 8066能广泛中和大量B型和C型HIV - 1病毒，而Fab 8062则无中和作用。晶体结构揭示了复合物中CDR - H2环构象的重要差异，这些差异延伸到抗原 - 抗体界面的其他区域，并表明中和特性和对靶标的亲和力至少部分可归因于CDR - H2环与抗原相互作用的差异。此外，对一个N - HR三聚体与三个Fab片段复合物的建模表明，CDR - H2环可能参与相邻抗体分子之间的紧密分子间接触，并且这种接触可能会阻碍N - HR三聚体与多于一个抗体分子形成复合物，这取决于由其与抗原相互作用所确定的结合的CDR - H2环的构象。与5 - 螺旋和另一种中和性单克隆抗体D5（通过完全不同的抗体文库和淘选程序获得）的复合物晶体结构进行比较，揭示了CDR - H2环在最佳序列和构象上的显著趋同。
HIV - 1与靶细胞的膜融合是病毒感染的第一步。这个过程涉及两种病毒包膜糖蛋白gp120和gp41在gp120与靶细胞膜上的CD4受体和趋化因子辅助受体结合之后的一系列构象变化。在融合过程中，以三聚体卷曲螺旋形式存在的gp41的保守N - 七肽重复序列（N - HR）是可及的，并为产生广泛中和抗体提供了一个有吸引力的靶点。在此我们展示了两种单克隆Fab片段与N - HR三聚体模拟物形成的复合物的晶体结构。这些Fab片段是从一个包含超过10¹⁰种人源特异性的合成人源组合抗体文库中获得的，首先通过对N - HR模拟物进行淘选，然后通过对CDR - H2互补决定区进行靶向多样化来实现亲和力成熟。其中一种Fab片段能广泛中和大量B型和C型HIV - 1的原代分离株，而另一种则无中和作用。我们的结构揭示了CDR - H2环在抗原识别中的关键作用以及它与HIV - 1中和特性的相关性。
The conserved internal trimeric coiled-coil of the N-heptad repeat (N-HR) of HIV-1 gp41 is transiently exposed during the fusion process by forming a pre-hairpin intermediate, thus representing an attractive target for the design of fusion inhibitors and neutralizing antibodies. In previous studies we reported a series of broadly neutralizing mini-antibodies derived from a synthetic naïve human combinatorial antibody library by panning against a mimetic of the trimeric N-HR coiled coil, followed by affinity maturation using targeted diversification of the CDR-H2 loop. Here we report crystal structures of the N-HR mimetic 5-Helix with two Fabs that represent the extremes of this series: Fab 8066 is broadly neutralizing across a wide panel of B and C type HIV-1 viruses, whereas Fab 8062 is non-neutralizing. The crystal structures reveal important differences in the conformations of the CDR-H2 loops in the complexes that propagate into other regions of the antigen-antibody interface, and suggest that both neutralization properties and affinity for the target can be attributed, at least in part, to the differences in the interactions of the CDR-H2 loops with the antigen. Furthermore, modeling of the complex of an N-HR trimer with three Fabs suggests that the CDR-H2 loop may be involved in close intermolecular contacts between neighboring antibody molecules, and that such contacts may hinder the formation of complexes between the N-HR trimer and more than one antibody molecule depending on the conformation of the bound CDR-H2 loop which is defined by its interactions with antigen. Comparison with the crystal structure of the complex of 5-Helix with another neutralizing monoclonal antibody known as D5, derived using an entirely different antibody library and panning procedure, reveals remarkable convergence in the optimal sequence and conformation of the CDR-H2 loop. Membrane fusion of HIV-1 with its target cells represents the first step in viral infection. This process involves a series of conformational changes in two viral envelope glycoproteins, gp120 and gp41, subsequent to binding of gp120 to the CD4 receptor and the chemokine coreceptor on the target cell membrane. During the fusion process, the conserved N-heptad repeat (N-HR) of gp41 in the form of a trimeric coiled-coil is accessible and presents an attractive target for the generation of broadly neutralizing antibodies. Here we present the crystal structures of two monoclonal Fabs complexed to a mimetic of the N-HR trimer. These Fabs were derived from a synthetic human combinatorial antibody library comprising more than 1010 human specificities by first panning against an N-HR mimetic, followed by affinity maturation through targeted diversification of the CDR-H2 complementarity determining region. One of the Fabs is broadly neutralizing across a wide range of primary isolates from subtype B and C HIV-1, whereas the other one is non-neutralizing. Our structures reveal the key role of the CDR-H2 loop in antigen recognition and how this correlates with HIV-1 neutralization properties.
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影响因子：
64.5
作者：
Eckert, DM;Malashkevich, VN;Kim, PS
通讯作者：
Kim, PS
DOI：
10.1107/s0907444904019158
发表时间：
2004-12-01
影响因子：
2.2
作者：
Emsley, P;Cowtan, K
通讯作者：
Cowtan, K
影响因子：
5.6
作者：
CHOTHIA, C;LESK, AM
通讯作者：
LESK, AM
影响因子：
5.4
作者：
Gustchina, Elena;Louis, John M.;Clore, G. Marius
通讯作者：
Clore, G. Marius
影响因子：
7.3
作者：
Gustchina, E;Hummer, G;Clore, GM
通讯作者：
Clore, GM