Abstract
Chronic hepatitis B virus (HBV) poses a significant public health burden worldwide, encouraging the search for curative antivirals. One approach is capsid assembly modulators (CAMs), which are assembly agonists. CAMs lead to empty and defective capsids, inhibiting the formation of new viruses, and can also lead to defects in the release of the viral genome, inhibiting new infections. In this study, we employed hydrogen-deuterium exchange mass spectrometry (HDX-MS) to assess the impact of one such CAM, HAP18, on HBV dimers, capsids composed of 120 (or 90) capsid protein dimers, and cross-linked capsids (xl-capsids). HDX analysis revealed hydrogen bonding networks within and between the dimers. HAP18 disrupted the hydrogen bonding network of dimers, demonstrating a previously unappreciated impact on the dimer structure. Conversely, HAP18 stabilized both unmodified and cross-linked capsids. Intriguingly, cross-linking the capsid, which was accomplished by forming disulfides between an engineered C-terminal cysteine, increased the overall rate of HDX. Moreover, HAP18 binding induced conformational changes beyond the binding sites. Our findings provide evidence for allosteric communication within and between capsid protein dimers. These results show that CAMs are capable of harnessing this allosteric network to modulate the dimer and capsid dynamics.