Department of Physiology and Cellular Biophysics, Columbia University, New York, NY 10032, USA; National Resource for Automated Molecular Microscopy, Simons Electron Microscopy Center, New York Structural Biology Center, New York, NY 10027, USA.
Department of Microbiology, University of Alabama at Birmingham, Birmingham, AL 35294, USA.
Mol Cell. 2020 May 21;78(4):683-699.e11. doi: 10.1016/j.molcel.2020.04.014. Epub 2020 May 7.
Mycobacterium tuberculosis causes tuberculosis, a disease that kills over 1 million people each year. Its cell envelope is a common antibiotic target and has a unique structure due, in part, to two lipidated polysaccharides-arabinogalactan and lipoarabinomannan. Arabinofuranosyltransferase D (AftD) is an essential enzyme involved in assembling these glycolipids. We present the 2.9-Å resolution structure of M. abscessus AftD, determined by single-particle cryo-electron microscopy. AftD has a conserved GT-C glycosyltransferase fold and three carbohydrate-binding modules. Glycan array analysis shows that AftD binds complex arabinose glycans. Additionally, AftD is non-covalently complexed with an acyl carrier protein (ACP). 3.4- and 3.5-Å structures of a mutant with impaired ACP binding reveal a conformational change, suggesting that ACP may regulate AftD function. Mutagenesis experiments using a conditional knockout constructed in M. smegmatis confirm the essentiality of the putative active site and the ACP binding for AftD function.
结核分枝杆菌会引起结核病，这种疾病每年导致超过 100 万人死亡。它的细胞壁是常见的抗生素靶标，由于部分原因，其结构具有独特性，部分原因在于存在两种脂化多糖——阿拉伯半乳聚糖和脂阿拉伯甘露聚糖。阿拉伯呋喃糖基转移酶 D（AftD）是参与组装这些糖脂的必需酶。我们通过单颗粒冷冻电子显微镜确定了脓肿分枝杆菌 AftD 的 2.9 Å 分辨率结构。AftD 具有保守的 GT-C 糖基转移酶折叠和三个碳水化合物结合模块。糖基阵列分析表明，AftD 结合复杂的阿拉伯糖聚糖。此外，AftD 与酰基载体蛋白（ACP）非共价复合。与 ACP 结合能力受损的突变体的 3.4 和 3.5 Å 结构揭示了构象变化，表明 ACP 可能调节 AftD 的功能。使用在耻垢分枝杆菌中构建的条件性敲除进行的突变实验证实了假定的活性位点和 ACP 结合对 AftD 功能的重要性。
