Department of Chemistry, Université du Québec à Montréal, P.O. Box 8888, Downtown Station, Montreal H3C 3P8, Canada; Institut des Sciences de la Mer de Rimouski, Université du Québec à Rimouski, 310 allée des Ursulines, Rimouski G5L 3A1, Canada.
Department of Chemistry, Université du Québec à Montréal, P.O. Box 8888, Downtown Station, Montreal H3C 3P8, Canada.
Biochim Biophys Acta Biomembr. 2019 Apr 1;1861(4):871-878. doi: 10.1016/j.bbamem.2019.01.018. Epub 2019 Feb 2.
Vibrio splendidus is a marine bacterium often considered as a threat in aquaculture hatcheries where it is responsible for mass mortality events, notably of bivalves' larvae. This bacterium is highly adapted to dynamic salty ecosystems where it has become an opportunistic and resistant species. To characterize their membranes as a first and necessary step toward studying bacterial interactions with diverse molecules, we established a labelling protocol for in vivoH solid-state nuclear magnetic resonance (SS-NMR) analysis of V. splendidus. H SS-NMR is a useful tool to study the organization and dynamics of phospholipids at the molecular level, and its application to intact bacteria is further advantageous as it allows probing acyl chains in their natural environment and study membrane interactions. In this study, we showed that V. splendidus can be labelled using deuterated palmitic acid, and demonstrated the importance of surfactant choice in the labelling protocol. Moreover, we assessed the impact of lipid deuteration on the general fitness of the bacteria, as well as the saturated-to-unsaturated fatty acid chains ratio and its impact on the membrane properties. We further characterize the evolution of V. splendidus membrane fluidity during different growth stages and relate it to fatty acid chain composition. Our results show larger membrane fluidity during the stationary growth phase compared to the exponential growth phase under labelling conditions - an information to take into account for future in vivo SS-NMR studies. Our lipid deuteration protocol optimized for V. splendidus is likely applicable other microorganisms for in vivo NMR studies.
灿烂弧菌是一种海洋细菌，常被认为是水产养殖孵化场的威胁，因为它会导致大规模的贝类幼虫死亡事件。这种细菌高度适应动态咸水生境，已经成为一种机会主义和耐药的物种。为了研究其膜作为研究细菌与多种分子相互作用的第一步，我们建立了一种用于体内 H 固体核磁共振（SS-NMR）分析灿烂弧菌的标记方案。H SS-NMR 是研究磷脂在分子水平上的组织和动力学的有用工具，其应用于完整细菌具有进一步的优势，因为它可以在其天然环境中探测酰基链，并研究膜相互作用。在这项研究中，我们表明，灿烂弧菌可以用氘化棕榈酸进行标记，并证明了在标记方案中选择表面活性剂的重要性。此外，我们评估了脂质氘代对细菌一般适应性的影响，以及饱和脂肪酸与不饱和脂肪酸链的比例及其对膜性质的影响。我们进一步描述了在不同生长阶段中灿烂弧菌膜流动性的演变，并将其与脂肪酸链组成相关联。我们的结果表明，在标记条件下，与指数生长阶段相比，在静止生长阶段的膜流动性更大 - 这是未来体内 SS-NMR 研究中需要考虑的信息。我们为灿烂弧菌优化的脂质氘代方案可能适用于其他微生物的体内 NMR 研究。
