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Biochim Biophys Acta Biomembr. 2019 Apr 1;1861(4):871-878. doi: 10.1016/j.bbamem.2019.01.018. Epub 2019 Feb 2.

Labelling strategy and membrane characterization of marine bacteria Vibrio splendidus by in vivo2H NMR.

Author information

1
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.
2
Department of Chemistry, Université du Québec à Montréal, P.O. Box 8888, Downtown Station, Montreal H3C 3P8, Canada.
3
Department of Chemistry, Université du Québec à Montréal, P.O. Box 8888, Downtown Station, Montreal H3C 3P8, Canada; UMR 7099, CNRS - Université Paris Diderot, IBPC, 13 rue Pierre et Marie Curie, F-75005 Paris, France.
4
Institut des Sciences de la Mer de Rimouski, Université du Québec à Rimouski, 310 allée des Ursulines, Rimouski G5L 3A1, Canada.
5
Department of Chemistry, Université du Québec à Montréal, P.O. Box 8888, Downtown Station, Montreal H3C 3P8, Canada. Electronic address: marcotte.isabelle@uqam.ca.

Abstract

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 vivo2H solid-state nuclear magnetic resonance (SS-NMR) analysis of V. splendidus. 2H 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.

KEYWORDS:

Deuterium; Fatty acids; In-cell NMR; Isotopic labelling; Magic-angle spinning; Membrane fluidity

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