Evaluation of peptide-aldehyde inhibitors using R188I mutant of SARS 3CL protease as a proteolysis-resistant mutant

Akaji, Kenichi; Konno, Hiroyuki; Onozuka, Mari; Makino, Ayumi; Saito, Hiroyuki; Nosaka, Kazuto

The 3C-like (3CL) protease of the severe acute respiratory syndrome (SARS) coronavirus is a key enzyme for the virus maturation. We found for the first time that the mature SARS 3CL protease is subject to degradation at 188Arg/189Gln. Replacing Arg with Ile at position 188 rendered the protease resistant to proteolysis. The R188I mutant digested a conserved undecapeptide substrate with a K(m) of 33.8 μM and k(cat) of 4753 s(−1). Compared with the value reported for the mature protease containing a C-terminal His-tag, the relative activity of the mutant was nearly 10(6). Novel peptide-aldehyde derivatives containing a side-chain-protected C-terminal Gln efficiently inhibited the catalytic activity of the R188I mutant. The results indicated for the first time that the tetrapeptide sequence is enough for inhibitory activities of peptide-aldehyde derivatives.