Vaccinia virus A46 is an anti-inflammatory and non-anti-apoptotic, two-domain member of the poxviral Bcl-2-like protein family that inhibits the cellular innate immune response at the level of the Toll/interleukin-1 receptor (TIR) domain-containing TLR adaptor proteins MAL, MyD88, TRAM, and TRIF. The mechanism of interaction of A46 with its targets has remained unclear. The TIR domains of MAL and MyD88 have been shown to signal by forming filamentous assemblies. We show a clear concentration-dependent destruction of both of these assemblies by A46 by means of negative-stain electron microscopy from molar ratios of 1:15 for MAL and 1:30 for MyD88. Using targeted mutagenesis and protein-protein crosslinking, we show that A46 interacts with MAL and MyD88 through several facets, including residues on helices α1 and α7 and the C-terminal flexible region. We propose a model in which A46 targets the MAL and MyD88 signalosome intra-strand interfaces and gradually destroys their assemblies in a concentration-dependent manner.

痘苗病毒 A46 是痘病毒 Bcl-2 样蛋白家族的抗炎和非抗凋亡的双结构域成员，可在 Toll/白细胞介素-1 受体 (TIR) 水平抑制细胞先天免疫反应包含域的 TLR 衔接蛋白 MAL、MyD88、TRAM 和 TRIF。A46 与其目标相互作用的机制尚不清楚。MAL 和 MyD88 的 TIR 域已显示通过形成丝状组件来发出信号。我们通过负染色电子显微镜显示了 A46 对这两个组件的明显浓度依赖性破坏，MAL 的摩尔比为 1:15，MyD88 的摩尔比为 1:30。使用靶向诱变和蛋白质-蛋白质交联，我们表明 A46 通过几个方面与 MAL 和 MyD88 相互作用，包括螺旋 α1 和 α7 上的残基以及 C 端柔性区域。我们提出了一个模型，其中 A46 以 MAL 和 MyD88 信号小体链内接口为目标，并以浓度依赖的方式逐渐破坏它们的程序集。