Characterization of lipopolysaccharide from a heptoseless mutant of Escherichia coli by carbon 13 nuclear magnetic resonance.
Lipopolysaccharide (LPS) isolated from Escherichia coli D31m4, a heptoseless mutant, was studied by 13C and 31P NMR spectroscopy. Modified isolation and purification procedures are described which permitted high resolution NMR spectra to be obtained from samples of intact LPS. 31P NMR was used to monitor the purity and native heterogeneity of LPS samples. The anomeric carbon region of the 13C NMR spectrum taken at pH 7 contained five resonances that were assigned on the basis of chemical shift correlation, 13C-1H couplings, and T1 relaxation times. Two resonances, at 99.9 and 100.8 ppm, were attributed to two residues of 3-deoxy-D-manno-octulosonate (KDO) of which both were tentatively assigned to the alpha configuration. The Lipid A moiety gave rise to resonances at 94.0 and 94.9 ppm, both assigned to GlcNI, and a resonance at 103.1 ppm, assigned to GlcNII. The two anomeric carbon resonances observed for GlcNI reflected the variable substitution of C-1 with monophosphate or diphosphate groups. GlcNI and GlcNII were ascertained to be of the alpha and beta anomeric configuration, respectively, through chemical shift comparisons with model saccharides. The accepted KDO linkage site at C-3' of GlcNII appears not to be supported by the 13C chemical shift data.
D002241:Carbohydrates / Q000032:analysis; D004926:Escherichia coli / Q000235:genetics*; D006539:Heptoses / Q000235:genetics*; D008070:Lipopolysaccharides / Q000302:isolation & purification*; D009682:Magnetic Resonance Spectroscopy; D009154:Mutation
D002241:Carbohydrates; D006539:Heptoses; D008070:Lipopolysaccharides