Bordetella bronchiseptica, known to infect animals and rarely humans, expresses a lipopolysaccharide that plays an essential role in host interactions, being critical for early clearance of the bacteria. On a B. bronchiseptica 9.73 isolate, mutants defective in the expression of genes involved in the biosynthesis of the core region were previously constructed. Herein, a comparative detailed structural analysis of the expressed lipids A by MALDI-TOF mass spectrometry was performed. The Bb3394 LPS defective in a 2-amino-2-deoxy-d-galacturonic acid lateral residue of the core presented a penta-acylated diglucosamine backbone modified with two glucosamine phosphates, similar to the wild-type lipid A. In contrast, BbLP39, resulting in the interruption of the LPS core oligosaccharide synthesis, presented lipid A species consisting in a diglucosamine backbone N-substituted with C14:0(3-O-C12:0) in C-2 and C14:0(3-O-C14:0) in C-2 ', O-acylated with C14:0(3-O-C10:0(3-OH) in C-3 ' and with a pyrophosphate in C-1. Regarding Bb3398 also presenting a rough LPS, the lipid A is formed by a hexa-acylated diglucosamine backbone carrying one pyrophosphate group in C-1 and one phosphate in C-4 ', both substituted with ethanolamine groups. As far as we know, this is the first description of a phosphoethanolamine modification in B. bronchiseptica lipid A. Our results demonstrate that although gene deletions were not directed to the lipid A moiety, each mutant presented different modifications. MALDI-TOF mass spectrometry was an excellent tool to highlight the structural diversity of the lipid A structures biosynthesized during its transit through the periplasm to the final localization in the outer surface of the outer membrane.
Lipopolysaccharide, B.bronchiseptica, Lipid A modifications, UV-MALDI-TOF MS
Publication DOI: 10.1007/s13361-019-02233-3Journal NLM ID: 9010412Publisher: Elsevier
Correspondence: Alicia Couto
Institutions: Facultad de Ciencias Exactas y Naturales, Departamento de Quimica Organica - Consejo Nacional de Investigaciones Cientificas y Tecnicas, Centro de Investigacion en Hidratos de Carbono (CIHIDECAR), Universidad de Buenos Aires, Ciudad Universitaria, Intendente Guiraldes 2160, C1428GA, Buenos Aires, Argentina
Methods: SDS-PAGE, acid hydrolysis, MALDI-TOF MS, UV-MALDI-TOF MS, MALDI-LID-MS/MS
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