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bioRxiv · 10.64898/2026.01.27.701950

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

Zhong, Q.·Barritt, J. D.·Nji, E.·Gkeka, A.·Rouse, S. L.·Hohenester, E.·Tiengwe, C.
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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) alters antibody epitopes and influences infection outcome, but the responsible enzyme remained elusive. Here, we show that an expression-site-associated gene (ESAG3), which is co-transcribed with the active VSG, encodes an O-glucosyltransferase required for VSG3 O-glycosylation in vivo. ESAG3 depletion abolishes VSG3 O-glycosylation and recognition of live trypanosomes by an O-glycan-dependent monoclonal antibody, while complementation restores both. ESAG3 has strict UDP-glucose donor specificity and modifies serine/threonine residues within VSG3 peptides in vitro. Cryo-electron microscopy at 3.4 Angstrom resolution reveals that ESAG3 forms a homo-octadecameric ring with C3 symmetry, an architecture not previously observed among glycosyltransferases. Structure-guided mutagenesis shows that disruption of ESAG3 inter-subunit interfaces reduces higher-order assembly, enhances substrate conversion, and alters VSG glycoform composition. Furthermore, we show that VSG3 O-glycan occupancy correlates with susceptibility to complement-mediated trypanolysis in an antibody-dependent manner, establishing that ESAG3-mediated O-glucosylation shapes both antibody specificity and the functional outcome of the host response.

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Zhong, Q., Barritt, J. D., Nji, E., Gkeka, A., Rouse, S. L., Hohenester, E., Tiengwe, C.. 2026-01-27. An octadecameric O-glucosyltransferase generates diversity in antibody epitopes on variant surface antigens in African trypanosomes. https://doi.org/10.64898/2026.01.27.701950

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