Following phospholipid transfer through the OmpF3-MlaA-MlaC lipid shuttle with native mass spectrometry

• 成果类型: Article
• 署名作者: Kirschbaum, Carla; Bennett, Jack L.; Tian, Qiaoyu; Sen, Navoneel; Smith, Iain P. S.; Wu, Di; Benesch, Justin L. P.; Khalid, Syma; Isom, Georgia; Robinson, Carol, V
• 署名单位: University of Oxford; University of Oxford; University of Oxford; University of Oxford
• 刊物名称: PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
• ISSN/ISSBN: 0027-11977
• DOI: 10.1073/pnas.2420041122
• 发表日期: 2025-04-08
• 关键词: escherichia-coli; asymmetry; complex; gui

摘要

The maintenance of lipid asymmetry (Mla) system in gram- negative bacteria transfers phospholipids between the outer and inner membrane to maintain the outer membrane asymmetry. Misplaced phospholipids are extracted from the outer leaflet of the outer membrane by MlaA, transferred to the periplasmic lipid transporter MlaC, and shuttled to the inner membrane. We set out to investigate the lipid transfer between MlaA and MlaC using native mass spectrometry, with the aim of determining the lipid preferences of MlaC and whether MlaA preselected lipids for MlaC. First, we characterized the lipids that copurified with overexpressed MlaC, phosphatidylglycerol (PG), and phosenrichment of cyclopropane lipids. Under native expression conditions, we found that PG is three- fold enriched on MlaC compared to its abundance in the membrane. Next, we isolated and characterized OmpF3-MlaA complexes and demonstrated their ability to enhance loading of delipidated MlaC with bacterial and nonbacterial phospholipids. We then captured the intact ternary lipid shuttle (OmpF3-MlaA-MlaC) and demonstrated results point to a high population of endogenous PG on periplasmic MlaC, which likely arises from disassembly of the lipid shuttle to maintain lipid asymmetry for cell viability.