Department of Biochemistry, Faculty of Natural Sciences, Comenius University in Bratislava, 842 15 Bratislava, Slovakia.
Institute of Chemistry, Slovak Academy of Sciences, 845 38 Bratislava, Slovakia.
Proc Natl Acad Sci U S A. 2021 Apr 27;118(17). doi: 10.1073/pnas.2023663118.
one of the deadliest pathogens in human history, is distinguished by a unique, multilayered cell wall, which offers the bacterium a high level of protection from the attacks of the host immune system. The primary structure of the cell wall core, composed of covalently linked peptidoglycan, branched heteropolysaccharide arabinogalactan, and mycolic acids, is well known, and numerous enzymes involved in the biosynthesis of its components are characterized. The cell wall biogenesis takes place at both cytoplasmic and periplasmic faces of the plasma membrane, and only recently some of the specific transport systems translocating the metabolic intermediates between these two compartments have been characterized [M. Jackson, C. M. Stevens, L. Zhang, H. I. Zgurskaya, M. Niederweis, , 10.1021/acs.chemrev.0c00869 (2020)]. In this work, we use CRISPR interference methodology in to functionally characterize an ATP-binding cassette (ABC) transporter involved in the translocation of galactan precursors across the plasma membrane. We show that genetic knockdown of the transmembrane subunit of the transporter results in severe morphological changes and the accumulation of an aberrantly long galactan precursor. Based on similarities with structures and functions of specific O-antigen ABC transporters of gram-negative bacteria [C. Whitfield, D. M. Williams, S. D. Kelly, 295, 10593-10609 (2020)], we propose a model for coupled synthesis and export of the galactan polymer precursor in mycobacteria.
它是人类历史上最致命的病原体之一，其独特的多层细胞壁使其能够免受宿主免疫系统的攻击。细胞壁核心的一级结构由共价连接的肽聚糖、支链杂多糖阿拉伯半乳聚糖和分枝菌酸组成，其结构已经明确，并且已经对参与其成分生物合成的许多酶进行了特征描述。细胞壁的生物发生发生在细胞质和质膜的周质面，直到最近，一些特定的运输系统才被鉴定出来，这些系统可以将代谢中间产物在这两个隔室之间转运[M. Jackson, C. M. Stevens, L. Zhang, H. I. Zgurskaya, M. Niederweis,, 10.1021/acs.chemrev.0c00869 (2020)]。在这项工作中，我们使用 CRISPR 干扰方法在 中对涉及半乳糖前体跨质膜转运的 ATP 结合盒（ABC）转运蛋白进行功能表征。我们表明，转运蛋白的跨膜亚基的基因敲低会导致严重的形态变化和异常长的半乳糖前体的积累。基于与革兰氏阴性细菌特定 O 抗原 ABC 转运蛋白的结构和功能的相似性[C. Whitfield, D. M. Williams, S. D. Kelly, 295, 10593-10609 (2020)]，我们提出了分枝杆菌中半乳糖聚合物前体的偶联合成和外排模型。
