蛋白水解级联包括几个重要的生理系统,包括称为补体级联的先天免疫的主要分支。为了防止补体介导的攻击,莱姆病的病原体,伯氏螺旋体,产生许多外表面定位的脂蛋白,有助于成功的补体逃避。最近,我们发现了一对OspEF相关蛋白家族的Bergdorferi表面脂蛋白,称为ElpB和ElpQ,它们抑制抗体介导的补体激活。在这项研究中,我们使用一系列生化和生物物理方法研究了ElpB和ElpQ补体抑制的分子机制。补体活化的体外测定显示,每个Elp蛋白的独立折叠的同源C-末端结构域保持完全的补体抑制活性并选择性地抑制经典途径。使用结合测定法和补体成分C1s酶测定法,我们显示Elp蛋白与活化的C1s的结合通过与C1s-C4结合竞争而不封闭活性位点来阻断补体成分C4的切割。C1s介导的C4裂解依赖于活化诱导的结合位点,称为exosite。为了测试这些外部位点是否参与Elp-C1s结合,我们进行了定点诱变,这表明ElpB和ElpQ结合需要位于C1s的丝氨酸蛋白酶结构域上的阴离子结合外位点中的C1s残基。基于这些结果,我们提出了一个模型,其中ElpB和ElpQ利用激活诱导的构象变化,这对于C1s介导的C4切割通常很重要。我们的研究扩展了已知的微生物病原体补体逃避机制,并揭示了莱姆病螺旋体选择性抑制C1s的新分子机制。
Proteolytic cascades comprise several important physiological systems, including a primary arm of innate immunity called the complement cascade. To safeguard against complement-mediated attack, the etiologic agent of Lyme disease, Borreliella burgdorferi, produces numerous outer surface-localized lipoproteins that contribute to successful complement evasion. Recently, we discovered a pair of B. burgdorferi surface lipoproteins of the OspEF-related protein family-termed ElpB and ElpQ-that inhibit antibody-mediated complement activation. In this study, we investigate the molecular mechanism of ElpB and ElpQ complement inhibition using an array of biochemical and biophysical approaches. In vitro assays of complement activation show that an independently folded homologous C-terminal domain of each Elp protein maintains full complement inhibitory activity and selectively inhibits the classical pathway. Using binding assays and complement component C1s enzyme assays, we show that binding of Elp proteins to activated C1s blocks complement component C4 cleavage by competing with C1s-C4 binding without occluding the active site. C1s-mediated C4 cleavage is dependent on activation-induced binding sites, termed exosites. To test whether these exosites are involved in Elp-C1s binding, we performed site-directed mutagenesis, which showed that ElpB and ElpQ binding require C1s residues in the anion-binding exosite located on the serine protease domain of C1s. Based on these results, we propose a model whereby ElpB and ElpQ exploit activation-induced conformational changes that are normally important for C1s-mediated C4 cleavage. Our study expands the known complement evasion mechanisms of microbial pathogens and reveals a novel molecular mechanism for selective C1s inhibition by Lyme disease spirochetes.