Antigenic and glycan-binding characteristics of norovirus GII.20 capsid protein VP1.

Human norovirus (NoV) is one of the major pathogens causing acute gastroenteritis. The GII.20 NoV VP1 reveals a close evolutionary relationship with that of the widely prevalent GII.4 genotype based on sequence phylogenetic analysis. In this study, we characterized the antigenic properties and glycan binding profile of GII.20 VP1 protein. GII.20 VP1 protein was successfully produced using a baculovirus expression system, and self-assembled into virus-like particles (VLPs) with a diameter of approximately 38 nm. Immunological assays demonstrated that polyclonal sera generated against GII.20 VLPs exhibited strong binding activity at dilution up to 1:8,192,000, while maintaining detectable cross-reactivity with GII.4 VLP at 1:128,000. dilution. Reciprocal cross-reactivity was observed, with anti-GII.4 sera recognizing GII.20 VLP at 1:125,000 dilution. Blocking assays demonstrated mutual partial cross-blocking between the antisera, with half-maximal blocking dilutions of 36 (anti-GII.20 sera) and 297 (anti-GII.4). Blocking assays demonstrated mutual partial cross-blocking effects between the antisera, with half-maximal blocking dilution of 36 and 297 for anti-GII.20 and anti-GII.4 polyclonal sera, respectively. Saliva binding analysis revealed that GII.20 VLPs recognize A, B, and O blood type saliva. Sequence alignment showed conserved key amino acids in the putative glycan-binding region between GII.20 and GII.4 genotypes. Structural modeling of the GII.20 P domain revealed high similarity with the corresponding domains of both GII.17 and GII.4 NoVs. These findings establish that GII.20 VP1 shares functional characteristics with GII.4 in both glycan binding specificity and antigenic cross-reactivity, providing more insights into the molecular epidemiology of diverse NoV genotypes.