Conformational dynamics of OMPs, periplasmic chaperones and BamA

OMP、周质伴侣和 BamA 的构象动力学

• 批准号: 421343516
• 负责人: Professor Dr. Michael Schlierf
• 金额: --
• 依托单位: B CUBE - Center for Molecular Bioengineering
• 依托单位国家: 德国
• 项目类别: Research Grants
• 资助国家: 德国
• 项目状态: 未结题
• 来源: https://gepris.dfg.de/gepris/projekt/421343516?language=en
• 关键词: Conformational; dynamics; OMPs; periplasmic; chaperones

Outer membrane proteins maintain the viability of Gram-negative bacteria as receptors, transporters, ion channels, lipases, and porins. Folding and assembly of outer membrane proteins involves synchronized action of periplasmic chaperones and multi-protein machineries, which escort the highly hydrophobic polypeptides to their target outer membrane in a folding competent state. Over the past decades proteins and their biochemical function along the biogenesis pathway have been identified. Yet, the mechanisms of action and the intriguing ability of all these molecular machines to work without the typical cellular energy source of ATP, but solely based on thermodynamic principles are not well understood. In this application we aim to understand specific interactions of the periplasmic chaperones Skp and SurA with large outer membrane proteins, specific recognition sites and synergies of both chaperones. We will monitor these dynamics and structures using time-resolved single-molecule FRET, which allows us to reveal kinetics from the nanosecond to the minute timescale and spatial information from 2 to 8 nanometer. We further aim to reveal insights on the lateral gate dynamics of BamA and how those dynamics are influenced by the lipidic environment through reconstitution in artificial lipid bilayers and by preserving near native lipid compositions. We will further study how the lateral gate is influenced by other BAM proteins, by antibiotics and during insertion of OMP proteins. of the environment, direct interactions with other BAM proteins or antibiotics, and during insertion of OMP proteins.

外膜蛋白作为受体、转运体、离子通道、脂肪酶和孔蛋白维持革兰氏阴性菌的生存能力。外膜蛋白的折叠和组装涉及质周伴侣和多蛋白机制的同步作用，这些机制将高度疏水的多肽以折叠态护送到目标外膜。在过去的几十年里，人们已经确定了蛋白质及其在生物发生途径中的生化功能。然而，所有这些分子机器的作用机制和令人感兴趣的能力，没有典型的细胞能量来源ATP，但仅仅基于热力学原理，还没有很好地理解。在这个应用中，我们的目标是了解质周伴侣Skp和SurA与大外膜蛋白的特定相互作用，特定的识别位点和两种伴侣的协同作用。我们将使用时间分辨的单分子FRET来监测这些动力学和结构，这使我们能够揭示从纳秒到分钟的时间尺度和2到8纳米的空间信息的动力学。我们进一步旨在揭示BamA的侧门动力学，以及这些动力学如何受到脂质环境的影响，通过在人工脂质双分子层中重构并保持接近天然的脂质成分。我们将进一步研究其他BAM蛋白、抗生素和OMP蛋白插入过程中对侧门的影响。与其他BAM蛋白或抗生素的直接相互作用，以及在插入OMP蛋白期间。