title: Characterization of lipid A acylation patterns in Francisella tularensis, Francisella novicida, and Francisella philomiragia using multiple-stage mass spectrometry and matrix-assisted laser desorption/ionization on an intermediate vacuum source linear ion trap.
abstract: Lipopolysaccharide (LPS) is a major component of the outer membrane of Gram-negative bacteria. The lipid A region of LPS stimulates the immune system in a structure-dependent manner. We have previously identified the two major lipid A species from Francisella tularensis as asymmetric tetraacylated structures containing four long acyl chains (16 and 18 carbons) and a single phosphate group that is partially modified by galactosamine (Phillips, N. J.; Schilling B.; McLendon, M. K.; Apicella, M. A.; Gibson, B. W. Infect. Immun. 2004, 72, 5340-5348). In the current study, we used matrix-assisted laser desorption/ionization on an intermediate vacuum source (vMALDI) coupled to a linear ion trap (LIT) mass spectrometer in multiple-stage mass fragmentation mode (MSn) to determine the structures of several minor and low abundant lipid A species present in F. tularensis, Francisella novicida, and Francisella philomiragia LPS that have not been previously characterized. Comprehensive vMALDI-MSn fragmentation studies allowed us to deduce the composition and the position of the fatty acid substituents within the lipid A moieties. Unexpectedly, most of these minor lipid A species consisted of multiple isobaric species with acyl chains of various lengths. Moreover, we found that a small portion of these lipid A species may be modified by the addition of a hexose or hexosamine sugar, in addition to the galactosamine that was previously identified. Overall, we found that MSn analysis on the vMALDI-LIT-MS platform was highly efficient and sensitive, allowing for thorough analysis of very minor lipid A species.
authors.affiliation: The Buck Institute for Age Research, Novato, California 94945, USA.
authors.forename: Birgit
authors.initials: B
authors.lastname: Schilling
authors.forename: Molly K
authors.initials: MK
authors.lastname: McLendon
authors.forename: Nancy J
authors.initials: NJ
authors.lastname: Phillips
authors.forename: Michael A
authors.initials: MA
authors.lastname: Apicella
authors.forename: Bradford W
authors.initials: BW
authors.lastname: Gibson
mesh_terms: D000215:Acylation; D005603:Francisella / Q000737:chemistry*; D005604:Francisella tularensis / Q000737:chemistry; D006595:Hexosamines; D006601:Hexoses; D008050:Lipid A / Q000737:chemistry*; D008070:Lipopolysaccharides; D015394:Molecular Structure; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D014618:Vacuum
chemical_list: D006595:Hexosamines; D006601:Hexoses; D008050:Lipid A; D008070:Lipopolysaccharides