BINDING MODE OF CA074, A SPECIFIC IRREVERSIBLE INHIBITOR, TO BOVINE CATHEPSIN-B AS DETERMINED BY X-RAY CRYSTAL ANALYSIS OF THE COMPLEX

The binding mode of CA074 rbamoyl-oxirane-2-carbonyl)-L-isoleucyl-L-pr oline] , a specific irreversible inhibitor, to bovine spleen cathepsin B was elucidated by X-ray crystal structure analysis of the complex a t 2.2 Angstrom resolution (conventional R=0.185), Inconsistently with our model used for the development of CA074, the L-isoleucyl-L-proline and propylcarbamoyl moieties are located at the S' and S subsites res pectively, This unexpected binding is primarily due to (i) similar ext ended chain conformations (due to the same S configurations) at the ox irane C2 and C3 atoms of CA074 and (ii) the just fit formation of doub le hydrogen bonds between the carboxyl oxygens of L-proline and the im idazole nitrogens of His-110 and His-ill residues (these residues are missing in papain, the tertiary structure of which was used for the de sign of CA074), The oxirane C3 atom possessing the P' substituent is c ovalently bound to the Cys-29 S gamma atom (C3-S gamma=1.79 Angstrom) and the S configuration is maintained. The present result will provide useful information for characterizing the substrate-specificity of ca thepsin B.