Structure-based drug design on SARS protease and glutaminyl cyclase targets.
Andrew Wang

Severe acute respiratory syndrome (SARS) is an emerging infectious disease caused by a novel human coronavirus. In order to develop effective tools to combat the diseases associated with SARS and other coronaviruses, we need to understand the structure and function of their viral proteins. We have focused on the SARS main protease (3CLpro) which is required to cleave the virus-encoded polyproteins during viral maturation. We have solved the crystal structures of the wild-type and mutant SARS main protease as well as its complexes with several inhibitors. The dimeric structure of the C145A mutant protease shows that the active site of one protomer binds with the C-terminal six amino acids of the protomer from another asymmetric unit, mimicking the product-bound form and suggesting a possible mechanism for maturation and clues for drug design (Hsu et al., J. Biol. Chem. 280, 20762-74, 2005). We have developed a potent SARS coronavirus (CoV) 3CL protease inhibitor (TG-0205221), distinctly different from AG7088, which displayed remarkable activity against both SARS CoV and human CoV 229E replications. The crystal structure of TG-0205221 bound to SARS CoV 3CL protease has revealed a unique and unsymmetrical binding mode. Detailed comparisons of enzyme bindings between this inhibitor and a natural peptide substrate suggest future design of inhibitors of other viral proteases.
N-terminal pyroglutamate (pGlu) formation from its glutaminyl (or glutamyl) precursor is required in the maturation of numerous bioactive peptides. The aberrant formation of pGlu may be related to several pathological processes, such as osteoporosis and amyloidotic diseases. The pGlu formation is catalyzed by the enzyme glutaminyl cyclase (QC). To probe this important but poorly understood modification, we determined the structure of human QC in free form and bound to a substrate and three imidazole-derived inhibitors. The structure reveals an / scaffold akin to that of two-zinc exopeptidases but with several insertions and deletions, particularly in the active-site region. The relatively closed active site displays Two alternate conformations due to the different indole orientations of Trp-207 result in two substrate (glutamine t-butyl ester)-binding modes. The single zinc ion in the active site is coordinated to three conserved residues and one water molecule, which is replaced by an imidazole nitrogen upon binding of the inhibitors. Together with structural and kinetic analyses of several active-site-mutant enzymes, a catalysis mechanism of the formation of protein N-terminal pGlu is proposed. Our results provide a structural basis for the rational design of inhibitors against QC-associated disorders (Huang et al., Proc. Natl. Acad, Sci., 102, 13117-22, 2005).