Novel Structural Features of Human Norovirus Capsid

ABSTRACT Human noroviruses are a major cause of gastroenteritis, yet there are still no vaccines or antivirals available. Nevertheless, a number of vaccine candidates that are currently in clinical trials are composed of norovirus virus-like particles (VLPs). These VLPs are recognized as morphologically and antigenically similar to norovirus virions. An X-ray crystal structure of the prototype (GI.1) VLPs showed that the norovirus capsid has a T=3 icosahedral symmetry and is composed of 180 copies of the major capsid protein (VP1) that folds into three quasi-equivalent subunits (A, B, and C). In this study, we determined the cryo-EM structure of VLPs for two GII.4 noroviruses that were detected in 1974 and 2012. We showed that these VLPs had a T=4 symmetry and were composed of 240 copies of VP1. The VP1 on the T=4 VLPs adapted four quasi-equivalent subunits (termed A, B, C, and D), which formed two distinct dimers (A/B and C/D). We found that the T=4 protruding domain was elevated ~21 Å off the capsid shell, which was ~7 Å more than the previously determined for the T=3 GII.10 norovirus. Another interesting feature of the T=4 VLPs was a small cavity and flaplike structure located at the twofold axis. This structural feature was associated with the shell domain (D subunit) and disrupted the contiguous shell. Altogether, we showed that the T=4 VLPs had a number of structural similarities and differences with other noroviruses, but how these structural changes associate with norovirus virions could be important for vaccine studies. IMPORTANCE The discovery that the GII.4 VLPs (identified in 1974 and 2012, termed CHDC-1974 and NSW-2012, respectively) have a T=4 symmetry is of major significance, since the NSW-2012 is clinically important and previous structural and biochemical studies assumed noroviruses have a T=3 symmetry and are composed of 180 copies of VP1. More importantly, NSW-2012 norovirus shared 96% amino acid identity with a GII.4 vaccine candidate and our data suggests that this vaccine might also have a T=4 symmetry. Although it is not clear if the T=4 VLPs were an artifact of the insect cell expression system, the T=4 VLP vaccines might not recognize equivalent epitopes on T=3 virions, which will be important for future neutralization studies. Finally, further studies with other norovirus genotypes and virions are clearly needed in order to determine the level of this structural diversity.