Kuklewicz Jeremi, Zimmer Jochen
Department of Molecular Physiology and Biological Physics, University of Virginia School of Medicine, Charlottesville, VA, USA.
Howard Hughes Medical Institute, University of Virginia, Charlottesville, VA, USA.
Nature. 2024 Apr;628(8009):901-909. doi: 10.1038/s41586-024-07248-9. Epub 2024 Apr 3.
Capsular polysaccharides (CPSs) fortify the cell boundaries of many commensal and pathogenic bacteria. Through the ABC-transporter-dependent biosynthesis pathway, CPSs are synthesized intracellularly on a lipid anchor and secreted across the cell envelope by the KpsMT ABC transporter associated with the KpsE and KpsD subunits. Here we use structural and functional studies to uncover crucial steps of CPS secretion in Gram-negative bacteria. We show that KpsMT has broad substrate specificity and is sufficient for the translocation of CPSs across the inner bacterial membrane, and we determine the cell surface organization and localization of CPSs using super-resolution fluorescence microscopy. Cryo-electron microscopy analyses of the KpsMT-KpsE complex in six different states reveal a KpsE-encaged ABC transporter, rigid-body conformational rearrangements of KpsMT during ATP hydrolysis and recognition of a glycolipid inside a membrane-exposed electropositive canyon. In vivo CPS secretion assays underscore the functional importance of canyon-lining basic residues. Combined, our analyses suggest a molecular model of CPS secretion by ABC transporters.
荚膜多糖（CPSs）强化了许多共生菌和病原菌的细胞边界。通过依赖ABC转运蛋白的生物合成途径，CPSs在细胞内由脂质锚定物合成，并通过与KpsE和KpsD亚基相关的KpsMT ABC转运蛋白分泌穿过细胞膜。在这里，我们利用结构和功能研究揭示革兰氏阴性菌中CPS分泌的关键步骤。我们表明KpsMT具有广泛的底物特异性，足以使CPSs穿过细菌内膜，并且我们使用超分辨率荧光显微镜确定了CPSs的细胞表面组织和定位。对处于六种不同状态的KpsMT-KpsE复合物进行的冷冻电子显微镜分析揭示了一个被KpsE包裹的ABC转运蛋白、ATP水解过程中KpsMT的刚体构象重排以及在膜暴露的正电峡谷内对糖脂的识别。体内CPS分泌试验强调了峡谷内衬碱性残基的功能重要性。综合起来，我们的分析提出了ABC转运蛋白分泌CPS的分子模型。
