An octadecameric O-glucosyltransferase generates diversity in antibody epitopes on variant surface antigens in African trypanosomes

Authors

Zhong, Q., Barritt, J. D., Nji, E., Gkeka, A., Rouse, S. L., Hohenester, E., Tiengwe, C.

Abstract

Immune evasion in many pathogens relies on sequence variation to generate antigenic diversity. African trypanosomes use an additional strategy where O-glucosylation of variant surface glycoproteins (VSGs) generates heterogeneous glycans that alter antibody epitope recognition, influencing infection outcome. However, the VSG O-glycosylation enzyme has remained unknown. Here, we identify ESAG3 as the glycosyltransferase required for generating these O-glycan-dependent epitopes. ESAG3 depletion in vivo abolishes O-glycosylation-specific monoclonal antibody recognition, while complementation restores binding. ESAG3 has strict UDP-glucose specificity, manganese dependence, and modifies serine/threonine residues within cysteine-flanked VSG peptides. Single-particle cryo-electron microscopy reveals that ESAG3 forms an unprecedented octadecameric architecture with C3 symmetry at 3.4 angstrom resolution, a novel quaternary organisation for a glycosyltransferase. Structure-guided mutagenesis demonstrates essential active-site residues for catalysis, while interface mutations disrupt octadecamer assembly and enhance substrate turnover, indicating that oligomeric architecture regulates catalytic output. This work establishes ESAG3 as a kinetoplastid-specific glucosyltransferase and reveals the molecular basis whereby VSG O-glycosylation generates epitope diversity alongside sequence-based antigenic variation.

Preprint server: bioRxiv
The authors list and abstract were imported from bioRxiv on 29 Jan 2026.

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