Abstract
Noroviruses are the major cause of viral gastroenteritis and re-emerge worldwide every year, with GII.4 currently being the most frequent human genotype. The norovirus capsid protein VP1 is essential for host immune response. The P domain mediates cell attachment via histo blood-group antigens (HBGAs) in a strain-dependent manner but how these glycan-interactions actually relate to cell entry remains unclear. Here, hydrogen/deuterium exchange mass spectrometry (HDX-MS) is used to investigate glycan-induced protein dynamics in P dimers of different strains, which exhibit high structural similarity but different prevalence in humans. While the almost identical strains GII.4 Saga and GII.4 MI001 share glycan-induced dynamics, the dynamics differ in the emerging GII.17 Kawasaki 308 and rare GII.10 Vietnam 026 strain. The structural aspects of glycan binding to fully deamidated GII.4 P dimers have been investigated before. However, considering the high specificity and half-life of N373D under physiological conditions, large fractions of partially deamidated virions with potentially altered dynamics in their P domains are likely to occur. Therefore, we also examined glycan binding to partially deamidated GII.4 Saga and GII.4 MI001 P dimers. Such mixed species exhibit increased exposure to solvent in the P dimer upon glycan binding as opposed to pure wildtype. Furthermore, deamidated P dimers display increased flexibility and a monomeric subpopulation. Our results indicate that glycan binding induces strain-dependent structural dynamics, which are further altered by N373 deamidation, and hence hint at a complex role of deamidation in modulating glycan-mediated cell attachment in GII.4 strains.
Highlights
MI001 P dimers stored for 1 year at pH 7.3 and 5 ◦ C, a fraction of approximately 64% was deamidated at N373
MI001 P dimer, no other region was protected from HDX under fucose treatment, but increased deuteration in the main peak distribution was observed in the P2 domain of both genogroup II (GII).4 strains, suggesting a more exposed conformation (Figure 4D)
We address the differences in structural responses to glycan binding of norovirus P dimers of the Asian epidemic strain GII.17
Summary
Several biophysical techniques including nuclear magnetic resonance (NMR), X-ray crystallography, native mass spectrometry (native MS) and hydrogen-deuterium exchange mass spectrometry (HDX-MS) have been applied to characterize binding of P dimers and VLPs to HBGAs and other glycans [11,12,13,14,15,16,17,18,19,20] These studies revealed that glycan preferences and binding affinities are strongly genotype- and strain-dependent. Saga P dimers that strongly attenuates glycan binding This deamidation appears to be site specific and occurs in GII.. As this exchange strongly depends on solvent accessibility and hydrogen bonding patterns, the method can provide information about regions involved in ligand binding as well as changes in protein dynamics in solution [23] This makes it a valuable technique for identification of glycan induced structural dynamics in different strains as well as elucidation of altered protein dynamics in deamidated P dimers. The surface representation shows the glycan binding cleft in top-view (pdb 4z4s)
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