Department of Virology, University of Helsinki and Helsinki University Hospital, Helsinki, Finland.
Department of Biological and Environmental Science, University of Jyväskylä, Jyväskylä, Finland.
PLoS Pathog. 2021 Nov 15;17(11):e1009728. doi: 10.1371/journal.ppat.1009728. eCollection 2021 Nov.
The accessory protein Nef of human and simian immunodeficiency viruses (HIV and SIV) is an important pathogenicity factor known to interact with cellular protein kinases and other signaling proteins. A canonical SH3 domain binding motif in Nef is required for most of these interactions. For example, HIV-1 Nef activates the tyrosine kinase Hck by tightly binding to its SH3 domain. An archetypal contact between a negatively charged SH3 residue and a highly conserved arginine in Nef (Arg77) plays a key role here. Combining structural analyses with functional assays, we here show that Nef proteins have also developed a distinct structural strategy-termed the "R-clamp"-that favors the formation of this salt bridge via buttressing Arg77. Comparison of evolutionarily diverse Nef proteins revealed that several distinct R-clamps have evolved that are functionally equivalent but differ in the side chain compositions of Nef residues 83 and 120. Whereas a similar R-clamp design is shared by Nef proteins of HIV-1 groups M, O, and P, as well as SIVgor, the Nef proteins of SIV from the Eastern chimpanzee subspecies (SIVcpzP.t.s.) exclusively utilize another type of R-clamp. By contrast, SIV of Central chimpanzees (SIVcpzP.t.t.) and HIV-1 group N strains show more heterogenous R-clamp design principles, including a non-functional evolutionary intermediate of the aforementioned two classes. These data add to our understanding of the structural basis of SH3 binding and kinase deregulation by Nef, and provide an interesting example of primate lentiviral protein evolution.
人类和猿免疫缺陷病毒（HIV 和 SIV）的辅助蛋白 Nef 是一种重要的致病性因子，已知其与细胞蛋白激酶和其他信号蛋白相互作用。Nef 中存在一个典型的 SH3 结构域结合基序，这是其与大多数这些相互作用所必需的。例如，HIV-1 Nef 通过与 Hck 的 SH3 结构域紧密结合来激活其酪氨酸激酶活性。在 Nef 中，带负电荷的 SH3 残基与高度保守的精氨酸（Arg77）之间的典型接触在此过程中发挥了关键作用。通过结构分析与功能测定相结合，我们发现 Nef 蛋白还采用了一种独特的结构策略，称为“R 夹”，该策略有利于通过支撑 Arg77 形成这种盐桥。对进化上多样化的 Nef 蛋白的比较表明，已经进化出几种不同的 R 夹，它们在功能上等效，但 Nef 残基 83 和 120 的侧链组成不同。虽然 HIV-1 组 M、O 和 P 以及 SIVgor 的 Nef 蛋白共享类似的 R 夹设计，但来自东部黑猩猩亚种（SIVcpzP.t.s.）的 SIV 的 Nef 蛋白则专门使用另一种类型的 R 夹。相比之下，中非黑猩猩（SIVcpzP.t.t.）和 HIV-1 组 N 株的 SIV 表现出更多的 R 夹设计原则的异质性，包括上述两类的非功能性进化中间体。这些数据增加了我们对 Nef 与 SH3 结合和激酶失调控的结构基础的理解，并为灵长类慢病毒蛋白进化提供了一个有趣的例子。
