Chemical structure and immunoreactivity of the lipopolysaccharide of the deep rough mutant I-69 Rd-/b+ of Haemophilus influenzae.

From the lipopolysaccharide of the deep rough mutant I-69 Rd--/b+ of Haemophilus influenzae two oligosaccharides were obtained after de-O-acylation and separation by high-performance anion exchange chromatography. Their chemical structures were determined by one- and two-dimensional 1H-, 13C- and 31P-NMR spectroscopy as alphaKdo-4P-(2-->6)-betaGlcN-4P-(1-->6)-alphaGlcN-1P and alphaKdo-5P-(2-->6)-betaGlcN-4P-(1-->6)-alphaGlcN-1P. The specificity of mAbs S42-21 and S42-16 specific for Kdo-4P or Kdo-5P, respectively [Rozalski, A., Brade L., Kosma P., Moxon R., Kusumoto S., & Brade H. (1997). Mol. Microbiol. 23, 569--577] was confirmed with neoglycoconjugates obtained by conjugation of the isolated oligosaccharides to BSA. In addition, a mAb S42-10-8 with unknown epitope specificity could be assigned using the neoglycoconjugates described herein. This mAb binds to an epitope composed of the bisphosphorylated glucosamine backbone of lipid A and Kdo-4P, whereby the latter determines the specificity strictly by the position of the phosphate group.