Pattern recognition molecules (PRMs) form an important part of innate immunity, where they facilitate the response to infections and damage by triggering processes such as inflammation. The pentraxin family of soluble PRMs comprises long and short pentraxins, with the former containing unique N-terminal regions unrelated to other proteins or each other. No complete high-resolution structural information exists about long pentraxins, unlike the short pentraxins, where there is an abundance of both X-ray and cryoelectron microscopy (cryo-EM)-derived structures. This study presents a high-resolution structure of the prototypical long pentraxin, PTX3. Cryo-EM yielded a 2.5-Å map of the C-terminal pentraxin domains that revealed a radically different quaternary structure compared to other pentraxins, comprising a glycosylated D4 symmetrical octameric complex stabilized by an extensive disulfide network. The cryo-EM map indicated α-helices that extended N terminal of the pentraxin domains that were not fully resolved. AlphaFold was used to predict the remaining N-terminal structure of the octameric PTX3 complex, revealing two long tetrameric coiled coils with two hinge regions, which was validated using classification of cryo-EM two-dimensional averages. The resulting hybrid cryo-EM/AlphaFold structure allowed mapping of ligand binding sites, such as C1q and fibroblast growth factor-2, as well as rationalization of previous biochemical data. Given the relevance of PTX3 in conditions ranging from COVID-19 prognosis, cancer progression, and female infertility, this structure could be used to inform the understanding and rational design of therapies for these disorders and processes.

中文翻译：

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使用混合低温电子显微镜和 AlphaFold 方法确定 PTX3 结构提供了对配体结合和补体激活的深入了解

模式识别分子 (PRM) 是先天免疫的重要组成部分，它们通过触发炎症等过程促进对感染和损伤的反应。可溶性 PRM 的五聚蛋白家族包括长五聚蛋白和短五聚蛋白，前者包含与其他蛋白质或彼此无关的独特 N 末端区域。与短五聚糖不同，长五聚糖不存在完整的高分辨率结构信息，短五聚糖存在丰富的 X 射线和低温电子显微镜 (cryo-EM) 衍生结构。本研究展示了原型长正五聚糖 PTX3 的高分辨率结构。Cryo-EM 产生了 C 末端五聚糖结构域的 2.5-Å 图，揭示了与其他五聚糖相比完全不同的四级结构，包含由广泛的二硫化物网络稳定的糖基化 D4 对称八聚体复合物。cryo-EM 图表明 α-螺旋延伸未完全解析的五聚蛋白结构域的 N 末端。AlphaFold 用于预测八聚体 PTX3 复合物的剩余 N 末端结构，揭示了两个具有两个铰链区域的长四聚体卷曲螺旋，使用低温 EM 二维平均值的分类对其进行了验证。由此产生的混合 cryo-EM/AlphaFold 结构允许绘制配体结合位点，例如 C1q 和成纤维细胞生长因子-2，以及之前生化数据的合理化。鉴于 PTX3 在 COVID-19 预后、癌症进展和女性不育等疾病中的相关性，