In Mycobacterium tuberculosis, Rv3806c is a membrane-bound phosphoribosyltransferase (PRTase) involved in cell wall precursor production. It catalyses pentosyl phosphate transfer from phosphoribosyl pyrophosphate to decaprenyl phosphate, to generate 5-phospho-β-ribosyl-1-phosphoryldecaprenol. Despite Rv3806c being an attractive drug target, structural and molecular mechanistic insight into this PRTase is lacking. Here we report cryogenic electron microscopy structures for Rv3806c in the donor- and acceptor-bound states. In a lipidic environment, Rv3806c is trimeric, creating a UbiA-like fold. Each protomer forms two helical bundles, which, alongside the bound lipids, are required for PRTase activity in vitro. Mutational and functional analyses reveal that decaprenyl phosphate and phosphoribosyl pyrophosphate bind the intramembrane and extramembrane cavities of Rv3806c, respectively, in a distinct manner to that of UbiA superfamily enzymes. Our data suggest a model for Rv3806c-catalysed phosphoribose transfer through an inverting mechanism. These findings provide a structural basis for cell wall precursor biosynthesis that could have potential for anti-tuberculosis drug development.

在结核分枝杆菌中，Rv3806c 是一种参与细胞壁前体生成的膜结合磷酸核糖基转移酶 (PRTase)。它催化磷酸戊糖基从磷酸核糖焦磷酸转移到磷酸十异丙烯基，生成5-磷酸-β-核糖基-1-磷酸十异丙烯醇。尽管 Rv3806c 是一个有吸引力的药物靶点，但缺乏对该 PRTase 的结构和分子机制的了解。在这里，我们报告了 Rv3806c 在供体和受体结合状态下的低温电子显微镜结构。在脂质环境中，Rv3806c 是三聚体，形成类似 UbiA 的折叠。每个原聚体形成两个螺旋束，它们与结合的脂质一起，是体外 PRTase 活性所必需的。突变和功能分析表明，磷酸十异丙烯酯和焦磷酸核糖酯分别以与 UbiA 超家族酶不同的方式结合 Rv3806c 的膜内和膜外空腔。我们的数据提出了 Rv3806c 催化磷酸核糖通过反转机制转移的模型。这些发现为细胞壁前体生物合成提供了结构基础，可能具有抗结核药物开发的潜力。