Structural basis for catalysis and ubiquitin recognition by the Severe acute respiratory syndrome coronavirus papain‐like protease

Abstract

Papain‐like protease (PL ^pro) is one of two cysteine proteases involved in the proteolytic processing of the polyproteins of Severe acute respiratory syndrome coronavirus (SARS‐CoV). PL ^pro also shows significant in vitro deubiquitinating and de‐ISGylating activities, although the detailed mechanism is still unclear. Here, the crystal structure of SARS‐CoV PL ^pro C112S mutant in complex with ubiquitin (Ub) is reported at 1.4 Å resolution. The Ub core makes mostly hydrophilic interactions with PL ^pro, while the Leu‐Arg‐Gly‐Gly C‐terminus of Ub is located in the catalytic cleft of PL ^pro, mimicking the P4–P1 residues and providing the first atomic insights into its catalysis. One of the O atoms of the C‐terminal Gly residue of Ub is located in the oxyanion hole consisting of the main‐chain amides of residues 112 and 113. Mutations of residues in the PL ^pro–Ub interface lead to reduced catalytic activity, confirming their importance for Ub binding and/or catalysis. The structure also revealed an N‐cyclohexyl‐2‐aminethanesulfonic acid molecule near the catalytic triad, and kinetic studies suggest that this binding site is also used by other PL ^pro inhibitors. Overall, the structure provides a foundation for understanding the molecular basis of coronaviral PL ^pro catalysis.