Complete Structural Elucidation of a Novel Lipooligosaccharide from the Outer Membrane of the Marine Bacterium Shewanella pacifica

Shewanella pacifica is a Gram-negative microrganism that is able to grow in sea water. A novel lipooligosaccharide (LOS) has been isolated from the outer membrane of this bacterium and its primary structure fully characterised. For the first time, the presence of a 2,3-dihydroxypropanoic acid residue (glyceric acid) has been identified in the core region. The complete structure of the LOS was determined by compositional and methylation analyses, by MALDI mass spectrometry, and by 1H, 13C and 31P NMR spectroscopy on the oligosaccharides formed by selective degradation of the LOS. Strong alkaline treatment, aimed at recovering and identifying the complete carbohydrate backbone, was carried out by hydrazinolysis followed by de-N-acylation with hot KOH, whereas mild hydrazinolysis (de-O-acylation) allowed us to gain information about the nature of the phosphate and other non-carbohydrate substituents on the core oligosaccharide. Mild acid hydrolysis was employed to obtain a lipid A moiety with which further degradation and mass spectrometry experiments were carried out in order to determine its primary structure.