Deubiquitinase Activities of Papain-like Protease from Severe Acute Respiratory Syndrome ( SARS ) Coronavirus against Ubiquitin-like Proteins

SARS coronavirus papain-like protease ( PLpro) is responsible for the proteolytic processing of pp1a ( 1ab) amino terminus to release nsp1,nsp2 and nsp3,and plays an important regulatory role in 第 7 期 陈晓娟等: SARS 冠状病毒 PLpro 蛋白酶对泛素样分子的 DUB 活性 the assembling of SARS coronavirus replicase complex and the escape from host innate immunity. SARS PLpro has been reported of in vitro deubiquitinase ( DUB) activity,but the characteristics and functions of the DUB activity are poorly understood. In this study,we first constructed several PLpro mutation constructs with N-terminal Ubl domain or C-terminal transmembrane ( TM ) domain deletion and the PLpro mutants at its catalytic sites ( Cys-His-Asp ) , then assayed for the DeISGylation and DeSUMOylating activites. We found that Ubl-deleted PLpro remained same DeISGylation activity in vivo as the wildtype suggesting the Ubl domain was dispensable. The DeISGylation of SARS-CoV PLpro was dependent on Cys-His catalytic residues,but little affected in the Asp mutation of the catalytic triad ( Cys-His-Asp) . The mutation of SARS PLpro core domain did not affect the SUMO-conjugated cellular proteins,but the transmembrane form of PLpro ( PLpro-TM ) induced dramatic reduction of SUMOconjugated cellular proteins. Such DeSUMOylating activity relied on intact catalytic residues,indicating that the DeSUMOylation activity of SARS PLpro required both TM-and catalytic residues. These results demonstrated that SARS PLpro might have both DeISGylating and DeSUMOylating activities in vivo.