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bioRxiv · 10.64898/2026.05.28.728500

Structural basis for recognition of a gonococcal lipooligosaccharide epitope by monoclonal antibody 2C7 informs vaccine and immunotherapeutic design

Tsagkarakou, A. S.·Nieto- Fabregat, F.·Masi, A. A.·Zhang, Y.·Fan, F.·Shaughnessy, J.·Rice, P. A.·Huang, X.·Ram, S.·Beernink, P. T.·Silipo, A.
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Abstract

Neisseria gonorrhoeae causes gonorrhea and poses a growing global health threat driven by antimicrobial resistance and the lack of an effective vaccine. The lipooligosaccharide (LOS) epitope recognized by monoclonal antibody (mAb) 2C7 is expressed by most clinical isolates, making it an attractive therapeutic target. To determine the molecular basis for mAb 2C7 recognition, we conducted structural studies of mAb 2C7 in complex with an octasaccharide derived from LOS. NMR epitope mapping demonstrated that mAb 2C7 contacts two of three glycan chains: the {beta}-chain (lactose) and the {gamma}-chain (N-acetylglucosamine). The 1.6-[A] resolution crystal structure of the Fab 2C7-octasaccharide complex revealed that these chains make extensive hydrogen bonds and hydrophobic contacts within a cleft of the Fab. Molecular dynamics (MD) simulations showed that the {beta} and {gamma} chains adopt stable, specific conformations, contrasted with transient interactions of the chain. Informed by these structural data, we synthesized a tetrasaccharide comprising the {beta} and {gamma} chains joined by heptose II. The tetrasaccharide conjugated to a carrier protein bound mAb 2C7 by ELISA and Western blotting. Isothermal titration calorimetry showed that the tetrasaccharide and octasaccharide bound mAb 2C7 with similar affinities, indicating that the smaller oligosaccharide retains the key binding determinants. Binding was enthalpically driven, consistent with the polar interactions observed crystallographically and by MD. NMR experiments with the tetrasaccharide confirmed interactions with all four residues, establishing it as the minimal 2C7 epitope. Together, these studies provide a structural framework for rational design of glycan-based vaccines and antibody therapeutics against antimicrobial-resistant N. gonorrhoeae.

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Tsagkarakou, A. S., Nieto- Fabregat, F., Masi, A. A., Zhang, Y., Fan, F., Shaughnessy, J., Rice, P. A., Huang, X., Ram, S., Beernink, P. T., Silipo, A.. 2026-05-29. Structural basis for recognition of a gonococcal lipooligosaccharide epitope by monoclonal antibody 2C7 informs vaccine and immunotherapeutic design. https://doi.org/10.64898/2026.05.28.728500

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