Top-Down Strategies for the Structural Elucidation of Intact Gram-negative Bacterial Endotoxins.
Re-modelling of lipopolysaccharides, which are the primary constituent of the outer cell membrane of Gram-negative bacteria, modulates pathogenesis and resistance to microbials. Reported herein is the characterization of intact Gram-negative bacterial lipooligosaccharides (LOS) via a new strategy utilizing online liquid chromatography (LC) coupled with ultraviolet photodissociation (UVPD) mass spectrometry. Compared to collision-based MS/MS methods, UVPD and UVPD/HCD promoted a greater array of cleavages within both the glycan and lipid moieties, including C-C, C-N, C-O cleavages in the acyl chains as well as glycosidic and cross-ring cleavages, thus providing the most far-reaching structural characterization of LOS. This LC-MS/MS strategy affords a robust analytical method to structurally characterize complex mixtures of bacterial endotoxins that maintains the integrity of the core oligosaccharide and lipid A domains of LOS, providing direct feedback about the cell envelope architectures and LOS modification strategies involved in resistance host innate immune defense.
Department of Chemistry, The University of Texas at Austin, 1 University Station A5300, Austin, TX, USA 78712.
John P
JP
O'Brien
The University of Texas at Austin, Department of Molecular Biosciences, 2506 Speedway A5000, Austin, TX, USA 78712.
Brittany D
BD
Needham
The University of Texas at Austin, Department of Molecular Biosciences, 2506 Speedway A5000, Austin, TX, USA 78712.
Dusty B
DB
Brown
The University of Texas at Austin, Department of Molecular Biosciences, 2506 Speedway A5000, Austin, TX, USA 78712.
M Stephen
MS
Trent
Department of Chemistry, The University of Texas at Austin, 1 University Station A5300, Austin, TX, USA 78712.
Jennifer S
JS
Brodbelt