Inhibitor Recognition Specificity of MERS-CoV Papain-like Protease May Differ from That of SARS-CoV

• Published in: ACS Chemical Biology 2015-06, Vol.10 (6), p.1456-1465
• Main Authors: Lee, Hyun, Lei, Hao, Santarsiero, Bernard D, Gatuz, Joseph L, Cao, Shuyi, Rice, Amy J, Patel, Kavankumar, Szypulinski, Michael Z, Ojeda, Isabel, Ghosh, Arun K, Johnson, Michael E
• Format: Article
• Language: English
• Subjects: Allosteric Regulation; Amino Acid Sequence; Antiviral Agents - chemistry; Catalytic Domain; Cysteine Endopeptidases - chemistry; Endopeptidases - chemistry; High-Throughput Screening Assays; Humans; Kinetics; Middle East Respiratory Syndrome Coronavirus - chemistry; Middle East Respiratory Syndrome Coronavirus - enzymology; Models, Molecular; Molecular Sequence Data; Protease Inhibitors - chemistry; Protein Binding; Protein Structure, Secondary; Recombinant Proteins - chemistry; SARS Virus - chemistry; SARS Virus - enzymology; Species Specificity; Surface Plasmon Resonance; Ubiquitin Thiolesterase - antagonists & inhibitors; Ubiquitin Thiolesterase - chemistry; Viral Proteins - antagonists & inhibitors; Viral Proteins - chemistry
• ISSN: 1554-8929; 1554-8937
• DOI: 10.1021/cb500917m

The Middle East Respiratory Syndrome coronavirus (MERS-CoV) papain-like protease (PLpro) blocking loop 2 (BL2) structure differs significantly from that of SARS-CoV PLpro, where it has been proven to play a crucial role in SARS-CoV PLpro inhibitor binding. Four SARS-CoV PLpro lead inhibitors were tested against MERS-CoV PLpro, none of which were effective against MERS-CoV PLpro. Structure and sequence alignments revealed that two residues, Y269 and Q270, responsible for inhibitor binding to SARS-CoV PLpro, were replaced by T274 and A275 in MERS-CoV PLpro, making critical binding interactions difficult to form for similar types of inhibitors. High-throughput screening (HTS) of 25 000 compounds against both PLpro enzymes identified a small fragment-like noncovalent dual inhibitor. Mode of inhibition studies by enzyme kinetics and competition surface plasmon resonance (SPR) analyses suggested that this compound acts as a competitive inhibitor with an IC50 of 6 μM against MERS-CoV PLpro, indicating that it binds to the active site, whereas it acts as an allosteric inhibitor against SARS-CoV PLpro with an IC50 of 11 μM. These results raised the possibility that inhibitor recognition specificity of MERS-CoV PLpro may differ from that of SARS-CoV PLpro. In addition, inhibitory activity of this compound was selective for SARS-CoV and MERS-CoV PLpro enzymes over two human homologues, the ubiquitin C-terminal hydrolases 1 and 3 (hUCH-L1 and hUCH-L3).