Structure of Senecavirus A 3C Protease Revealed the Cleavage Pattern of 3C Protease in Picornaviruses.

Senecavirus A (SVA) is an emerging picornavirus infecting porcine of all age groups and causing foot and mouth disease (FMD)-like symptoms. One of its key enzymes is the 3C protease (3C pro ), which is similar to other picornaviruses and essential for virus maturation by controlling polyprotein cleavage and RNA replication. In this study, we reported the crystal structure of SVA 3C pro at a resolution of 1.9 Å and a thorough structural comparison against all published picornavirus 3C pro structures. Using statistical and graphical visualization techniques, we also investigated the sequence specificity of the 3C pro . The structure revealed that SVA 3C pro adopted a typical chymotrypsin-like fold with the S1 subsite as the most conservative site among picornavirus 3C pro . The surface loop, A1-B1 hairpin, adopted a novel conformation in SVA 3C pro and formed a positively charged protrusion around S' subsites. Correspondingly, SVA scissile bonds preferred Asp rather than neutral amino acids at P3' and P4'. Moreover, SVA 3C pro showed a wide range tolerance to P4 residue volume (acceptable range: 67 Å 3 to 141 Å 3 ), such as aromatic side chain, in contrast to other picornaviruses. In summary, our results provided valuable information for understanding the cleavage pattern of 3C pro . IMPORTANCE Picornaviridae is a group of RNA viruses that harm both humans and livestock. 3C pro is an essential enzyme for picornavirus maturation, which makes it a promising target for antiviral drug development and a critical component for virus-like particle (VLP) production. However, the current challenge in the development of antiviral drugs and VLP vaccines includes the limited knowledge of how subsite structure determines the 3C pro cleavage pattern. Thus, an extensive comparative study of various picornaviral 3C pro was required. Here, we showed the 1.9 Å crystal structure of SVA 3C pro . The structure revealed similarities and differences in the substrate-binding groove among picornaviruses, providing new insights into the development of inhibitors and VLP.