bioRxiv2021
The murine norovirus (MNV) capsid protein is the target for various neutralizing antibodies binding to distal tips of its protruding (P)-domain. The bile acid glycochenodeoxycholic acid (GCDCA), an important co-factor for murine norovirus (MNV) infection, has recently been shown to induce conformational changes in surface-loops and a contraction of the virion. Here, we employ protein NMR experiments using stable isotope labeled MNV P-domains to shed light on underlying molecular mechanisms. We observe two separate sets of NMR resonance signals for P-domain monomers and dimers, permitting analysis of the corresponding exchange kinetics. Unlike human norovirus GII.4 P-dimers, which exhibit a half-life in the range of several days, MNV P-dimers are very short lived with a half-life of about 17 s. Addition of GCDCA shifts the equilibrium towards the dimeric form by tightly binding to the P-dimers. In MNV virions GCDCA-mediated stabilization of the dimeric arrangement of P-domains generates a more ordered state, which in turn may entropically assist capsid contraction. Numerous long-range chemical shift perturbations (CSPs) upon addition of GCDCA reflect allosteric conformational changes as a feature accompanying dimer stabilization. In particular, CSPs indicate rearrangement of the EF loop, a target for various neutralizing antibodies. Indeed, treating MNV virions with GCDCA prior to neutralizing antibody exposure abolishes neutralization. These findings advance our understanding of GCDCA-induced structural changes of MNV capsids and experimentally support an intriguing viral immune escape mechanism relying on GCDCA-triggered conformational changes of the P-dimer. Significance StatementThis study sheds light on the role of glycochenodeoxycholic acid (GCDCA) in promoting murine norovirus (MNV) infection and immune escape. Binding of GCDCA to the dimeric P-domain has been well characterized by crystallography and cryo EM studies, showing that upon GCDCA binding, a 90{degrees} rotation of the P-domain occurs, which results in its collapse onto the underlying shell of the virus. Our NMR experiments now reveal P-dimer stability as a new dimension of plasticity of MNV capsids and suggest that capsid contraction is entropically assisted. Conformational changes as a feature of P-dimer stabilization eliminate recognition by neutralizing antibodies, no longer being able to prevent infection. These findings highlight key differences between human and MNV capsid structures, promote our understanding of MNV infection on a molecular level, and reveal a novel immune escape mechanism.