Structural Elucidation of the Lipooligosaccharide from the Deep-Sea Bacterium Rheinheimera pacifica KMM 1406^T

The chemical structure of the lipooligosaccharide (LOS) from the marine bacterium Rheinheimera pacifica KMM 1406^T was investigated. Analysis by SDS-PAGE with silver staining revealed a low-molecular-weight banding pattern characteristic of R-type lipopolysaccharide (LOS). The fatty acid composition was determined by GC-MS, revealing mainly dodecanoic (12:0), 3-hydroxydodecanoic [12:0(3-OH)], and 3-hydroxytetradecanoic [14:0(3-OH)] acids. The oligosaccharide (OS) was obtained by complete deacylation of the LOS. Monosaccharide analysis indicated the presence of D-GlcN, D-Gal, D-Glc, and a phosphorylated Kdo residue. Using NMR spectroscopy, the structure of the OS was established as: α-D-Galp-(1→2)-α-D-Galp-(1→6)-α-D-Glcp-(1→5)-α-Kdop4P-(2→6)-β-D-GlcpN4P-(1→6)-α-D-GlcpN1P. Negative-ion MALDI-TOF mass spectrometry of the lipid A domain demonstrated the presence of bisphosphorylated penta-acylated and tetra-acylated molecular species. Tandem mass spectrometry determined the distribution of the acyl chains on the glucosamine disaccharide backbone. The LOS is characterized by a short carbohydrate chain and a low acylation degree of the lipid A domain.