Picornaviral 3C cysteine proteinases have a fold similar to chymotrypsin-like serine proteinases

Abstract

The picornavirus family includes several pathogens such as poliovirus, rhinovirus (the major cause of the common cold), hepatitis A virus and the foot-and-mouth disease virus. Picornaviral proteins are expressed by direct translation of the genomic RNA into a single, large polyprotein precursor. Proteolysis of the viral polyprotein into the mature proteins is assured by the viral 3C enzymes, which are cysteine proteinases. Here we report the X-ray crystal structure at 2.3 A resolution of the 3C proteinase from hepatitis A virus (HAV-3C). The overall architecture of HAV-3C reveals a fold resembling that of the chymotrypsin family of serine proteinases, which is consistent with earlier predictions. Catalytic residues include Cys 172 as nucleophile and His 44 as general base. The 3C cleavage specificity for glutamine residues is defined primarily by His 191. The overall structure suggests that an intermolecular (trans) cleavage releases 3C and that there is an active proteinase in the polyprotein.

A structural model of picornavirus leader proteinases based on papain and bleomycin hydrolase.

“…The 2A and 3C proteinases utilize a cysteine residue as the active site nucleophile but possess significant primary sequence and structural identity with chymotrypsin-like serine proteinases (Bazan & Fletterick, 1989 ;Gorbalenya et al, 1989 ;Allaire et al, 1994 ;Matthews et al, 1994). In contrast, the leader (L) proteinase of FMDV is a cysteine proteinase which has been postulated to operate in a similar fashion to papain-like enzymes (Gorbalenya et al, 1991).…”

Design and synthesis of irreversible inhibitors of foot-and-mouth disease virus 3C protease

“…The best inhibitor of the series has a benzoyl cap (12). A benzoyl cap is also found to improve inhibition when the P4 Pro residue is removed (13); this shorter peptide is more active than the corresponding acetylated version (8) although loss of the P4 Pro results in less potent inhibition overall. In the co-crystal structure of the enzyme with a peptide substrate, the P5 position is mostly solvent exposed and lacks significant enzyme contacts 15c , which is illustrated in Figure 1.…”

“…This makes the FMDV 3C pro enzyme a potential drug target for preventing viral replication, with no known cellular homologues being found in susceptible hosts. The structures of 3C pro enzymes from the related HRV 12 , HAV 13 and PV 14 have been determined, and we have solved the structure of FMDV 3C pro 15 , which may assist this process. Structurally, the enzyme is found to be a chymotrypsin-like cysteine protease 13, 15a, 15c .…”

Structure determination of Murine Norovirus NS6 proteases with C-terminal extensions designed to probe protease-substrate interactions

“…separated by a cleft that contains a Cys-His-Asp/Glu catalytic triad similar in arrangement to 58! the Ser-His-Asp triad characteristic of serine proteases (Allaire et al, 1994;Matthews et al, 59! 1994).…”