正在开发各种人类免疫缺陷病毒1型(HIV-1)基因型的可溶性重组天然样(NL)包膜糖蛋白(Env)三聚体,作为旨在诱导广泛中和抗体(bNAb)的疫苗候选物。原型设计,命名为BG505SOSIP.664,包含一个亚基间二硫键(SOS)以共价连接gp120和gp41胞外域(gp41ECTO)亚基和一个点取代,I559P(IP),进一步稳定GP41ECTO组件。如果没有SOS和IP更改,蛋白水解裂解的三聚体倾向于分解为其组成gp120和gp41ECTO亚基。我们展示,然而,即缺乏SOS和/或IP变化的NL三聚体可以以足以分析其抗原性和热稳定性的量被亲和纯化。总的来说,这些三聚体变体具有与完全稳定的SOSIP.664版本高度可比的性质。我们得出结论,SOS和IP变化的主要作用是在表达过程中和之后显着增加三聚体的稳定性,从而允许产生有用的量。然而,一旦三聚体被纯化,SOS和IP的变化对bNAb和其他表位的热稳定性和抗原性仅有微妙的影响。重要性正在开发基于HIV-1env基因的重组三聚体蛋白,用于人体疫苗试验。这些蛋白质的一个特征是它们对病毒颗粒上被中和抗体靶向的包膜糖蛋白结构的模拟,即,防止细胞被感染的抗体。正在探索的一个疫苗概念是,当作为免疫原递送时,重组三聚体可能能够引发病毒中和抗体。通常使用的设计称为SOSIP.664,该术语反映用于稳定三聚体并允许它们以实际有用的量生产的序列变化。这里,我们表明,这些稳定的变化作用,以增加三聚体产量在生产细胞内的生物合成过程中,但对纯化三聚体的性质几乎没有影响。
Soluble recombinant native-like (NL) envelope glycoprotein (Env) trimers of various human immunodeficiency virus type 1 (HIV-1) genotypes are being developed as vaccine candidates aimed at the induction of broadly neutralizing antibodies (bNAbs). The prototypic design, designated BG505 SOSIP.664, incorporates an intersubunit disulfide bond (SOS) to covalently link the gp120 and gp41 ectodomain (gp41ECTO) subunits and a point substitution, I559P (IP), to further stabilize the gp41ECTO components. Without the SOS and IP changes, proteolytically cleaved trimers tend to disintegrate into their constituent gp120 and gp41ECTO subunits. We show, however, that NL trimers lacking the SOS and/or IP change can be affinity purified in amounts sufficient for analyses of their antigenicity and thermal stability. In general, these trimer variants have properties highly comparable to those of the fully stabilized SOSIP.664 version. We conclude that the major effect of the SOS and IP changes is to substantially increase trimer stability during and after the expression process, thereby allowing useful amounts to be produced. However, once the trimers have been purified, the SOS and IP changes have only subtle impacts on thermostability and the antigenicity of bNAb and other epitopes.IMPORTANCE Recombinant trimeric proteins based on HIV-1 env genes are being developed for vaccine trials in humans. A feature of these proteins is their mimicry of the envelope glycoprotein structure on virus particles that is targeted by neutralizing antibodies, i.e., antibodies that prevent cells from becoming infected. One vaccine concept under exploration is that recombinant trimers may be able to elicit virus-neutralizing antibodies when delivered as immunogens. A commonly used design is designated SOSIP.664, a term reflecting the sequence changes that are used to stabilize the trimers and allow their production in practically useful amounts. Here, we show that these stabilizing changes act to increase trimer yield during the biosynthesis process within the producer cell but have little impact on the properties of purified trimers.