Mechanism for controlling the monomer–dimer conversion of SARS coronavirus main protease

Abstract

The Severe acute respiratory syndrome coronavirus (SARS-CoV) main protease (M(pro)) cleaves two virion polyproteins (pp1a and pp1ab); this essential process represents an attractive target for the development of anti-SARS drugs. The functional unit of M(pro) is a homodimer and each subunit contains a His41/Cys145 catalytic dyad. Large amounts of biochemical and structural information are available on M(pro); nevertheless, the mechanism by which monomeric M(pro) is converted into a dimer during maturation still remains poorly understood. Previous studies have suggested that a C-terminal residue, Arg298, interacts with Ser123 of the other monomer in the dimer, and mutation of Arg298 results in a monomeric structure with a collapsed substrate-binding pocket. Interestingly, the R298A mutant of M(pro) shows a reversible substrate-induced dimerization that is essential for catalysis. Here, the conformational change that occurs during substrate-induced dimerization is delineated by X-ray crystallography. A dimer with a mutual orientation of the monomers that differs from that of the wild-type protease is present in the asymmetric unit. The presence of a complete substrate-binding pocket and oxyanion hole in both protomers suggests that they are both catalytically active, while the two domain IIIs show minor reorganization. This structural information offers valuable insights into the molecular mechanism associated with substrate-induced dimerization and has important implications with respect to the maturation of the enzyme.

Targeting the Dimerization of the Main Protease of Coronaviruses: A Potential Broad-Spectrum Therapeutic Strategy

“…In 2013, Wu et al presented the crystal structure of R298A mutant of SARS-CoV M pro in the presence of a peptide substrate. 40 The R298A mutant undergoes a reversible substrate induced dimerization with minute changes in the relative position of the domain III of each monomer as compared to wt M pro . As indicated by active enzyme centrifugation (AEC) experiments, the kinetic parameters of the R298A mutant were identical with that of wt M pro .…”

Crystal structure of the papain-like protease of MERS coronavirus reveals unusual, potentially druggable active-site features

“…4D). The M pro is a cysteine protease comprising a chymotrypsin-like fold; in these enzymes, the oxyanion hole is formed by the main-chain amides of the catalytic cysteine (or serine) and of the penultimate residue (Taranto et al, 2008;Wu et al, 2013). The penultimate residue before the catalytic nucleophile in chymotrypsin-like proteases is absolutely conserved as a glycine, whereas in MERS-CoV PL pro , the conserved Asn109 resides at this position.…”

Characterization and Inhibition of the Main Protease of Severe Acute Respiratory Syndrome Coronavirus

“…From the crystal structure, the N-terminal residues 1-7 (N-finger) are buried in the dimer interface with numerous contacts with the domain II close to the active site of the other protomer ( Fig. 3) [10,18,25,26]. The first crystal structure of SARS-CoV 3CL pro is complexed with a substrate-like hexapeptidyl chloromethylketone (Cbz-Val-Asn-Ser-Thr-Leu-Gln-CMK) inhibitor, revealing the structural features of this protease [10].…”