Department of Chemistry, University of Michigan, Ann Arbor, Michigan.
Department of Microbiology and Immunology, University of Michigan Medical School, Ann Arbor, Michigan.
Biophys J. 2018 Jul 17;115(2):242-250. doi: 10.1016/j.bpj.2017.12.015. Epub 2018 Jan 12.
Bacteroides thetaiotaomicron (Bt) is a prominent member of the human gut microbiota with an extensive capacity for glycan harvest. This bacterium expresses a five-protein complex in the outer membrane, called the starch utilization system (Sus), which binds, degrades, and imports starch into the cell. Sus is a model system for the many glycan-targeting polysaccharide utilization loci found in Bt and other members of the Bacteroidetes phylum. Our previous work has shown that SusG, a lipidated amylase in the outer membrane, explores the entire cell surface but diffuses more slowly as it interacts with starch. Here, we use a combination of single-molecule tracking, super-resolution imaging, reverse genetics, and proteomics to show that SusE and SusF, two proteins that bind starch, are immobile on the cell surface even when other members of the system are knocked out and under multiple different growth conditions. This observation suggests a new paradigm for protein complex formation: binding proteins form immobile complexes that transiently associate with a mobile enzyme partner.
拟杆菌属（Bt）是人类肠道微生物群中的一个重要成员，具有广泛的聚糖收获能力。这种细菌在外膜表达一个由 5 种蛋白质组成的复合物，称为淀粉利用系统（Sus），该系统结合、降解和将淀粉导入细胞。Sus 是许多聚糖靶向多糖利用基因座的模型系统，这些基因座存在于 Bt 和其他拟杆菌门成员中。我们之前的工作表明，SusG，一种位于外膜中的脂化淀粉酶，在整个细胞表面进行探索，但随着与淀粉的相互作用，扩散速度会变慢。在这里，我们使用单分子追踪、超分辨率成像、反向遗传学和蛋白质组学的组合，表明即使在系统的其他成员被敲除和在多种不同的生长条件下，与淀粉结合的两种蛋白质 SusE 和 SusF 仍在细胞表面处于无运动状态。这一观察结果为蛋白质复合物的形成提出了一个新的范例：结合蛋白形成无运动复合物，这些复合物与移动的酶伴侣短暂地结合。
