Shanghai Institute for Advanced Immunochemical Studies, School of Life Science and Technology, ShanghaiTech University, Shanghai 201210, China.
School of Life Sciences, Tianjin University, Tianjin 300072, China.
Proc Natl Acad Sci U S A. 2023 Jun 6;120(23):e2302858120. doi: 10.1073/pnas.2302858120. Epub 2023 May 30.
Arabinogalactan (AG) is an essential cell wall component in mycobacterial species, including the deadly human pathogen . It plays a pivotal role in forming the rigid mycolyl-AG-peptidoglycan core for in vitro growth. AftA is a membrane-bound arabinosyltransferase and a key enzyme involved in AG biosynthesis which bridges the assembly of the arabinan chain to the galactan chain. It is known that AftA catalyzes the transfer of the first arabinofuranosyl residue from the donor decaprenyl-monophosphoryl-arabinose to the mature galactan chain (i.e., priming); however, the priming mechanism remains elusive. Herein, we report the cryo-EM structure of AftA. The detergent-embedded AftA assembles as a dimer with an interface maintained by both the transmembrane domain (TMD) and the soluble C-terminal domain (CTD) in the periplasm. The structure shows a conserved glycosyltransferase-C fold and two cavities converging at the active site. A metal ion participates in the interaction of TMD and CTD of each AftA molecule. Structural analyses combined with functional mutagenesis suggests a priming mechanism catalyzed by AftA in AG biosynthesis. Our data further provide a unique perspective into anti-TB drug discovery.
阿拉伯半乳聚糖（AG）是分枝杆菌属物种（包括致命的人类病原体）细胞壁的重要组成部分。它在体外生长中形成刚性的酰基阿拉伯甘露聚糖-AG-肽聚糖核心中发挥关键作用。AftA 是一种膜结合的阿拉伯糖基转移酶，是参与 AG 生物合成的关键酶，它将阿拉伯聚糖链的组装与半乳聚糖链连接起来。已知 AftA 催化从供体脱植基单磷酸阿拉伯糖到成熟半乳聚糖链的第一个阿拉伯呋喃糖基残基的转移（即引发）；然而，引发机制仍然难以捉摸。在此，我们报告了 AftA 的冷冻电镜结构。去污剂嵌入的 AftA 以二聚体的形式组装，其界面由跨膜域（TMD）和周质中的可溶性 C 末端域（CTD）共同维持。该结构显示出保守的糖基转移酶-C 折叠和两个在活性位点汇聚的腔。一个金属离子参与每个 AftA 分子的 TMD 和 CTD 的相互作用。结构分析结合功能突变表明，AftA 在 AG 生物合成中催化了一种引发机制。我们的数据进一步为抗结核药物发现提供了独特的视角。
