The N-finger Peptide Of SARS-CoV 3CL～(pro) Is Essential For The Enzyme Activity

In later of 2002, an atypical pneumonia, emerged in Guangdong province, China. The disease was named the severe acute respiratory syndrome (SARS) by WHO and a novel coronavirus, termed SARS-CoV, was rapidly identified as the etiological agent. Coronaviruses are enveloped, plus-strand RNA viruses that feature the largest RNA genomes currently known.The SARS -CoV main protease, which is a 33.8KD protease (also called 3C-like protease,3CL^pro), plays a pivotal role in mediating viral replication and transcription functions through extensive proteolytic processing of two replicase polyproteins,ppla and pplab. So it is essential for the life cycle of SARS-CoV and therefore represents a key anti-viral target.The SARS 3C-like protease consists of the N-finger and two distinct folds: namely the N-terminal chymotrypsin fold containing the domains I and II hosting the complete catalytic machinery and the C-terminal helical domain III. To understand the functional role of the N-finger (Ser-Gly-Phe-Arg-Lys-Met-Ala), the genes encoding 3CL protease (3CL^pro) of SARS virus and its mutant 3CL^pro(â–³_1-7) that truncated with N-finger were constructed and expressed in E.coli system . The 3CL^pro showed a proteolytic activity on the fluorogenic substrate containing the consensus auto-cleavage site of 3CL^pro. However, the 3CL^pro(â–³_1-7) mutant showed no enzymatic activity at the same conditions indicating that the N-finger peptide of 3CL^pro is essential for the enzymatic activity though it is not the enzyme reactive center. We also synthesized the N-finger peptide by solid-phase peptide synthesis method and the peptide showed partially inhibitory effect on the SARS 3CL protease. Base on the results above, a novel strategy is proposed to design drug for anti-SARS CoV.