| 1. |
- Dzieciatkowska, Monika, et al.
(författare)
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Characterization of intact lipopolysaccharides from the Haemophilus influenzae strain RM 118 using electrophoresis-assisted open-tubular liquid chromatography-mass spectrometry
- 2008
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Ingår i: Electrophoresis. - : Wiley. - 0173-0835 .- 1522-2683. ; 29:10, s. 2171-2181
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Tidskriftsartikel (refereegranskat)abstract
- We have applied an electrophoresis-assisted open-tubular LC-MS method for analyzing intact lipopolysaccharides (LPSs) from Haemophilus influenzae strain RM 118 (Rd). We were able to obtain structural information on both core oligosaccharides (OSs) and the lipid A moiety including the sialylation, glycylation, and the distribution of fatty acid residues on the disaccharide backbone of lipid A. The fragmentation patterns of sodiated and protonated LPS molecules were investigated for determining the location of sialic acid. It was found that the tandem mass spectra of sodiated ions provided unambiguous evidence of both sialylated lactose and sialylated lacto-N-neotetraose. In contrast, the fragment ions of protonated ions only offered the evidence for the existence of sialylated lacto-N-neotetraose. The lipid A of Gram-negative bacteria, as the principal endotoxic component of LP S, plays a major role in the pathogenesis of bacterial infections. We have previously characterized lipid. A species after mild acid hydrolysis of LPS during which lipid A precipitates. In this study, intact LPS was directly introduced to a tandem mass spectrometer. In-source dissociation strategy was employed, followed by multiple-stage MS/MS on the ions originating from the lipid part to obtain structural information. This is the first time that the structure of lipid A of H. influenzae was characterized by MS/MS on intact LPS molecules without any prior chemical modifications. In the same way information on the OS can be obtained by MS/MS by focusing on ions originating from core OS.
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| 2. |
- Dzieciatkowska, Monika, et al.
(författare)
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Rapid method for sensitive screening of oligosaccharide epitopes in the lipooligosaccharide from Campylobacter jejuni strains isolated from Guillain-Barre syndrome and Miller Fisher syndrome patients
- 2008
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Ingår i: Journal of Clinical Microbiology. - 0095-1137 .- 1098-660X. ; 46:10, s. 3429-3436
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Tidskriftsartikel (refereegranskat)abstract
- Campylobacter jejuni lipooligosaccharide (LOS) can trigger Guillain-Barre syndrome (GBS) due to its similarity to human gangliosides. Rapid and accurate structural elucidation of the LOS glycan of a strain isolated from a GBS patient could help physicians determine the spectrum of anti-ganglioside antibodies likely to be found and therefore provide valuable assistance in establishing an appropriate course of treatment. The ability of implemented mass spectrometry-based approaches in a clinical setting has been limited by the laborious and time-consuming nature of the protocols, typically 3 to 4 days, used to prepare LOS. In order to improve the analytical throughput, microwave-assisted enzymatic digestion was investigated. In this study, the bacterial cells were suspended in 50 mu l of 20 mM ammonium acetate buffer containing DNase and RNase and treated by direct microwave irradiation for 3 min. Then, proteinase K was added and the samples were again microwaved. The intact LOS samples were analyzed using electrophoresis-assisted open-tubular liquid chromatography-mass spectrometry. The reliability of the rapid, high-throughput technique was demonstrated through analysis of LOS glycans from 73 C. jejuni strains. The structure was elucidated using material from a single colony. The total time for sample preparation and MS analysis is less than 60 min.
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| 3. |
- Li, Jianjun, et al.
(författare)
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Structural characterization of sialylated glycoforms of H-influenzae by electrospray mass spectrometry : fragmentation of protonated and sodiated O-deacylated lipopolysaccharides
- 2007
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Ingår i: Rapid Communications in Mass Spectrometry. - : Wiley. - 0951-4198 .- 1097-0231. ; 21:6, s. 952-960
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Tidskriftsartikel (refereegranskat)abstract
- Sialylated lipopolysaccharide (LPS) glycoforms from Haemophilus influenzae were characterized by tandem mass spectrometry using a new generation hyphenated mass spectrometer which combines a triple quadrupole and a linear ion trap (Q-Trap). The fragmentation of both protonated and sodiated molecular ions from O-deacylated LPS (LPS-OH) obtained in MS2 experiments in the positive mode was studied. The MS2 spectra of protonated ions provided unambiguous evidence for the presence and sequence of sialylated lactosamine present in lacto-N-neotetraose oligosaccharide extensions but not for sialyl-lactose structures whilst fragmentation of sodiated adducts, [M+Na](+), afforded information diagnostic of mono- and disialylated lactose extensions. To study this we used a highly sialylated LPS from a H. influenzae strain capable of sialyl-lactose expression only. We then applied the method to the H. influenzae genome strain, Rd, in which glycoforms containing both sialyl-lactose and sialyl-lacto-N-neotetraose were detected from diagnostic B-ions at m/z 638.2 ([Neu5Ac(1) Hex(2)+ Na](+)) and 657.2 ([Neu5Ac(1) Hex(1) HexNAc(1)+H](+)). Unique fragmentation patterns provided the locations and sequences of these oligosaccharide extensions. This is the first time both sialylated lactose and sialylated lacto-N-neotetraose units have been detected and characterized by tandem mass spectrometry in the same molecule. This methodology is of general applicability for determination of common sialylated oligosaccharide extension in bacterial LPS.
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