Evolution Of The Interactions Between Gii.4 Noroviruses And Histo-Blood Group Antigens: Insights From Experimental And Computational Studies

Author summary Human norovirus (NoV) has been recognized as the leading cause of the epidemic acute gastroenteritis worldwide and more than 50% acute gastroenteritis outbreaks are associated with NoVs. NoVs are highly infectious and may result in serious dehydration, malnutrition and even death, which severely threatens human health and brings heavy economic burden. NoVs are highly genetically diverse, in which the GII.4 genotype is the most predominant. The reported outbreaks of NoV infections have risen sharply from 2002, and it is suggested that the increasing NoV infections are attributed to the emergence of new strains with more infectiousness. GII.4 NoV evolves rapidly and on average every 2-3 years a new strain appears. It has been revealed that the histo-blood group antigens (HBGAs) serve as the recognition receptor for the GII.4 NoVs infecting the host cell, and the NoV-HBGA interactions may play an important role in selecting the predominate variants during the evolution of GII.4 NoVs. However, the molecular mechanism behind the evolution of the NoV-HBGA binding affinities is still not clear. In this work, the representative GII.4 NoV strains prevalent in the past decades were expressed, and the changes in the interactions between these strains and the receptor HBGAs were investigated by using the experimental measurements combined with computational simulations. Based on the experimental and computational results, a molecular mechanism that accounts for the increasing of the NoV-HBGA binding affinities during the evolution of GII.4 NoVs was proposed. Our studies are helpful for the understanding of the evolution mechanism of GII.4 NoVs and provide valuable information for the drug and vaccine designs against GII.4 NoVs.Norovirus (NoV) is the major pathogen causing the outbreaks of the viral gastroenteritis across the world. Among the various genotypes of NoV, GII.4 is the most predominant over the past decades. GII.4 NoVs interact with the histo-blood group antigens (HBGAs) to invade the host cell, and it is believed that the receptor HBGAs may play important roles in selecting the predominate variants by the nature during the evolution of GII.4 NoVs. However, the evolution-induced changes in the HBGA-binding affinity for the GII.4 NoV variants and the mechanism behind the evolution of the NoV-HBGA interactions remain elusive. In the present work, the virus-like particles (VLPs) of the representative GII.4 NoV stains epidemic in the past decades were expressed by using the Hansenula polymorpha yeast expression platform constructed by our laboratory, and then the enzyme linked immunosorbent assay (ELISA)-based HBGA-binding assays as well as the molecular dynamics (MD) simulations combined with the molecular mechanics/generalized born surface area (MMGBSA) calculations were performed to investigate the interactions between various GII.4 strains and different types of HBGAs. The HBGA-binding assays show that for all the studied types of HBGAs, the evolution of GII.4 NoVs results in the increased NoV-HBGA binding affinities, where the early epidemic strains have the lower binding activity and the newly epidemic strains exhibit relative stronger binding intensity. Based on the MD simulation and MMGBSA calculation results, a physical mechanism that accounts for the increased HBGA-binding affinity was proposed. The evolution-involved residue mutations cause the conformational rearrangements of loop-2 (residues 390-396), which result in the narrowing of the receptor-binding pocket and thus tighten the binding of the receptor HBGAs. Our experimental and computational studies are helpful for better understanding the mechanism behind the evolution-induced increasing of HBGA-binding affinity, which may provide useful information for the drug and vaccine designs against GII.4 NoVs.