Structural basis for the dimerization and substrate recognition specificity of porcine epidemic diarrhea virus 3C-like protease

Porcine epidemic diarrhea virus (PEDV), a member of the genus Alphacoronavirus, has caused significant damage to the Asian and American pork industries. Coronavirus 3C-like protease (3CL^pro), which is involved in the processing of viral polyproteins for viral replication, is an appealing antiviral drug target. Here, we present the crystal structures of PEDV 3CL^pro and a molecular complex between an inactive PEDV 3CL^pro variant C144A bound to a peptide substrate. Structural characterization, mutagenesis and biochemical analysis reveal the substrate-binding pockets and the residues that comprise the active site of PEDV 3CL^pro. The dimerization of PEDV 3CL^pro is similar to that of other Alphacoronavirus 3CL^pros but has several differences from that of SARS-CoV 3CL^pro from the genus Betacoronavirus. Furthermore, the non-conserved motifs in the pockets cause different cleavage of substrate between PEDV and SARS-CoV 3CL^pros, which may provide new insights into the recognition of substrates by 3CL^pros in various coronavirus genera.