The Crystal Structure of N?-p-tosyl-lysyl Chloromethylketone-Bound Oligopeptidase B from Serratia Proteamaculans Revealed a New Type of Inhibitor Binding

A covalent serine protease inhibitor—Na-p-Tosyl-Lysyl Chloromethylketone (TCK) is a modified lysine residue tosylated at the N-terminus and chloromethylated C-terminus, one molecule of which capable forming two bonds with both Ser His catalytic residues, was co-crystallized oligopeptidase B (OpB) from Serratia proteomaculans (PSPmod). The kinetics study, preceded crystallization, shows that stoichiometry TCK-dependent inhibition PSPmod 1:2 (protein:inhibitor). crystal structure PSPmod-TCK complex, solved resolution 2.3 Å, confirmed new type inhibitor binding. Two TCK molecules were bound to enzyme molecule: Ser, other His. Due this mode binding, intermediate state disturbed conformation triad preserved in complex. Nevertheless, analysis amino acid surroundings its comparison antipain-bound OpB Trypanosoma brucei provided an insight substrate-binding pocket. Supposedly, binding typical for interaction chloromethylketone derivatives two-domain OpBs. In open conformational these enzymes are assumed solution, disordered configuration prevents simultaneous residues.