An essential saccharide binding domain for the mAb 2C7 established for Neisseria gonorrhoeae LOS by ES-MS and MSn

A study of bacterial surface oligosaccharides were investigated among diffe rent strains of Neisseria gonorrhoeae to correlate structural features esse ntial for binding to the MAb 2C7, This epitope is widely expressed and cons erved in gonococcal isolates, characteristics essential to an effective can didate vaccine antigen. Sample lipooligosaccharides (LOS), was prepared by a modification of the hot phenol-water method from which de-O-acetylated LO S and oligosaccharide (OS) components were analyzed by ES-MS-CID-MS and ES- MSn in a triple quadrupole and an ion trap mass spectrometer, respectively. Previously documented natural heterogeneity was apparent from both LOS and OS preparations which was admired with fragments induced by hydrazine and mild acid treatment. Natural heterogeneity was limited to phosphorylation a nd antenni extensions to the alpha-chain, Mild acid hydrolysis to release O S also hydrolyzed the beta(1-->6) glycosidic linkage of lipid A. OS structu res were determined by collisional and resonance excitation combined with M S and multistep MSn which provided sequence information from both neutral l oss, and nonreducing terminal fragments. A comparison of OS structures, wit h earlier knowledge of MAb binding, enzyme treatment, and partial acid hydr olysis indicates a generic overlapping domain for 2C7 binding. Reoccurring structural features include a Hep alpha(1-->3)Hep beta(l-->5)KDO trisacchar ide core branched on the nonreducing terminus (Hep-2) with an alpha(l-->2) linked GlcNAc (gamma-chain), and an alpha-linked lactose (beta-chain) resid ue. From the central heptose (Hep-1), a beta(1-->4) linked lactose (alpha-c hain), moiety is required although extensions to this residue appear unnece ssary.