Multivalency influences virus dynamics at the glycocalyx: A study of glycosaminoglycan binding in HPV16 entry

Virus attachment at the cell surface often involves the establishment of multiple ligand-receptor interactions between viral capsid proteins and glycans in the glycocalyx. Multivalency likely contributes to strengthening and fine-tuning virus dynamics at the cell surface to optimize entry. Here, we present experimental and theoretical frameworks to describe how virus attachment, detachment, and diffusion at the cell surface are modulated by multivalency. We focus on human papillomavirus 16 (HPV16), a leading cause of cervical cancer and its multivalent interactions with heparan sulfate (HS), a ubiquitous cell-surface glycan. Using single-particle tracking microscopy, we investigate the dynamic behavior of HPV16 particles interacting with multiple HS chains through a glycocalyx mimic consisting of end-tethered heparin chains that were selectively desulfated. We further establish a theoretical framework to predict the dynamic behavior of individual virus particles interacting with multiple cellular receptors and apply it to the HPV16-HS binding system using previously published association and dissociation rates of the individual capsomer-glycan bonds. Our experiments reveal that N-sulfation is essential to ensure HPV16 association and validate our theoretical prediction that at biologically relevant timescales, the lifetime of monovalent interaction primarily influences the apparent particle attachment behavior. Conversely, the nature of the sulfate group had a marginal effect on particle dissociation in the experiments, due to the great influence of multivalency on the particle's interaction lifetime. Experiments and simulations also indicate that the mobility of HPV16 particles on heparin surfaces is highly restricted due to the relatively high affinity of the monovalent interactions, suggesting that particle motion due to the creation and rupturing of individual bonds is unlikely key in HPV16 recruitment. Together, these findings provide quantitative mechanistic insights into how virus-glycan interactions are modulated. They highlight the importance of HS chemistry in modulating early HPV16 engagement and suggest new targets against viral binding.