Spike and host glycan determinants of HKU1 airway tropism

Creutznacher, R.; Wallace, L. E.; Debski-Antoniak, O. J.; Drulyte, I.; Hulswit, R. J. G.; Beekman, J.; van Kuppeveld, F. J. M.; Bosch, B.-J.; de Haan, C. A. M.; Frank, M.; Hurdiss, D. L.; de Groot, R. J.

doi:10.64898/2026.08.05.741156

bioRxiv · 10.64898/2026.08.05.741156

Spike and host glycan determinants of HKU1 airway tropism

Creutznacher, R.·Wallace, L. E.·Debski-Antoniak, O. J.·Drulyte, I.·Hulswit, R. J. G.·Beekman, J.·van Kuppeveld, F. J. M.·Bosch, B.-J.·de Haan, C. A. M.·Frank, M.·Hurdiss, D. L.·de Groot, R. J.

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Abstract

Human coronavirus HKU1 comprises two distinct serotypes, A and B, whose spike proteins are substantially divergent. Here, we show that spikes from both serotypes preferentially bind 9-O-acetylated 2,8-linked disialosides. Cryo-electron microscopy of the B-type N5 spike reveals a conserved extended binding site in domain S1A that accommodates both the terminal and penultimate sialic acid residues, with interactions involving the penultimate residue substantially enhancing binding. Spike N-glycan processing modulates affinity and linkage selectivity; glycans flanking the binding pocket offer a plausible structural basis for these effects. In contrast to the HKU1-A spike, which adopts open S1B-up conformations upon ligand binding, the N5 apo structure showed that the ligand-binding site was already formed and the e1 relay element register-shifted in most protomers. Nevertheless, we detected neither spontaneous opening nor a transition to an S1B-up state following ligand binding. These findings, obtained with a minimally modified ectodomain, differ from recent reports of ligand-independent opening. Molecular dynamics simulations indicated that membrane-embedded GT3, but not GD3, presents its glycan chain in a geometry compatible with S1A-mediated engagement. Concordantly, in human nasal epithelial cultures cell surface GT3-like O-acetylated trisialoside glycotopes were detected in ciliated cells, linking their cell-type-specific presentation to HKU1 tropism.

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Creutznacher, R., Wallace, L. E., Debski-Antoniak, O. J., Drulyte, I., Hulswit, R. J. G., Beekman, J., van Kuppeveld, F. J. M., Bosch, B.-J., de Haan, C. A. M., Frank, M., Hurdiss, D. L., de Groot, R. J.. 2026-08-05. Spike and host glycan determinants of HKU1 airway tropism. https://doi.org/10.64898/2026.08.05.741156

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