Format:
Online-Ressource
ISSN:
1522-2683
Content:
Abstract: We recently demonstrated that the combined use of lipopolysaccharide (LPS) reverse staining and high‐efficiency passive elution techniques can be successfully used as a suitable interface between LPS slab‐gel separation and electrospray ionization‐mass spectrometry (ESI‐MS) of LPS‐derived oligosaccharides. Here, we extend our micropurification strategy for the analysis of O‐deacylated LPS forms from Vibrio fischeri HMK after recovery from single reverse‐stained LPS bands using matrix‐assisted laser desorption/ionization‐time of flight‐mass spectrometry (MALDI‐TOF‐MS). The quantities (30–40 μg) obtained from the two gel‐resolved LPS bands were sufficient to allow MALDI‐TOF‐MS detection of O‐deacylated LPS glycoforms at m/z 3767.1, 3890.1 for the high‐molecular‐weight or at m/z 2522.5, 2645.4, 2725.7, and 2848.7 for the low‐molecular‐weight LPS band. These LPS band heterogeneities resulted not only from variations in the oligosaccharide region of the LPS but also from two phosphorylation states of the lipid A (diphosphoryl and diphosphoryl plus a single phosphoethanolamine substitution). On the other hand, MALDI‐TOF mass spectra of the separated LPS bands displayed reduced heterogeneity and increased signal‐to‐noise ratios as compared to spectra of the unpurified LPS. Furthermore, micropurification of LPS bands prior MALDI‐TOF‐MS led to a higher sensitivity of detection of less abundant low‐molecular‐weight LPS glycoforms. Taken together, this and our previous study on gel‐micropurified LPS using ESI definitively show how one can unambiguously determine the different molecular species contained within each gel‐separated LPS band, their relative abundance and oligosaccharide sequences.
In:
volume:25
In:
number:14
In:
year:2004
In:
pages:2156-2164
In:
extent:9
In:
Electrophoresis, Weinheim : Wiley-Blackwell, 1980-, 25, Heft 14 (2004), 2156-2164 (gesamt 9), 1522-2683
Language:
English
DOI:
10.1002/elps.200405980
URN:
urn:nbn:de:101:1-2023100307140611695467
URL:
https://doi.org/10.1002/elps.200405980
URL:
https://nbn-resolving.org/urn:nbn:de:101:1-2023100307140611695467
URL:
https://d-nb.info/1304796221/34
URL:
https://doi.org/10.1002/elps.200405980
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