School of Biological and Behavioural Sciences, Queen Mary, University of London, Mile End Road, London E1 4NS, UK; Cancer Research Horizons, Level 4NW The Francis Crick Institute, 1 Midland Road, London NW1 1AT, UK.
School of Biological and Behavioural Sciences, Queen Mary, University of London, Mile End Road, London E1 4NS, UK.
J Mol Biol. 2023 Aug 1;435(15):168146. doi: 10.1016/j.jmb.2023.168146. Epub 2023 May 17.
One infection method widely used by many gram-negative bacteria involves a protein nanomachine called the Type Three Secretion System (T3SS). The T3SS enables the transportation of bacterial "toxins" via a proteinaceous channel that directly links the cytosol of the bacteria and host cell. The channel from the bacteria is completed by a translocon pore formed by two proteins named the major and minor translocators. Prior to pore formation, the translocator proteins are bound to a small chaperone within the bacterial cytoplasm. This interaction is crucial to effective secretion. Here we investigated the specificity of the binding interfaces of the translocator-chaperone complexes from Pseudomonas aeruginosa via the selection of peptide and protein libraries based on its chaperone PcrH. Five libraries encompassing PcrH's N-terminal and central α-helices were panned, using ribosome display, against both the major (PopB) and minor (PopD) translocator. Both translocators were shown to significantly enrich a similar pattern of WT and non-WT sequences from the libraries. This highlighted key similarities/differences between the interactions of the major and minor translocators with their chaperone. Moreover, as the enriched non-WT sequences were specific to each translocator, it would suggest that PcrH can be adapted to bind each translocator individually. The ability to evolve such proteins indicates that these molecules may provide promising anti-bacterial candidates.
一种被许多革兰氏阴性菌广泛使用的感染方法涉及一种称为 III 型分泌系统（T3SS）的蛋白质纳米机器。T3SS 通过蛋白质通道将细菌“毒素”运送到细菌和宿主细胞的细胞质直接相连的地方。该通道由两个称为主要和次要转运蛋白的蛋白质组成的转位器孔完成。在形成孔之前，转运蛋白与细胞质内的小伴侣结合。这种相互作用对于有效的分泌至关重要。在这里，我们通过基于其伴侣蛋白 PcrH 的肽和蛋白质文库的选择，研究了铜绿假单胞菌的转运体-伴侣复合物的结合界面的特异性。使用核糖体展示技术，针对主要（PopB）和次要（PopD）转运体，对包含 PcrH 的 N 端和中央α-螺旋的五个文库进行了淘选。两种转运体均从文库中显著富集了相似的 WT 和非 WT 序列模式。这突出了主要和次要转运体与其伴侣蛋白之间相互作用的关键相似之处/差异。此外，由于富集的非 WT 序列与每个转运体特异性相关，这表明 PcrH 可以适应单独与每个转运体结合。这些蛋白质能够进化的能力表明，这些分子可能提供有前途的抗菌候选物。
