Substrate Specificity of Bovine Cathepsin B and Its Inhibition by CA 074, Based on Crystal Structure Refinement of the Complex

Abstract

The crystal structure of the bovine spleen cathepsin B (BSCB)-CA074 complex was refined to R=0.152 using X-ray diffraction data up to 2.18 Å resolution. BSCB is charac-terized by an extra Cys148-Cys252 disulfide bridge, as compared with rat and human CBs. Although the crystal structures of these enzymes showed similar overall folding, a difference was observed in the occluding loop, a structural element specific only to CB. Comparison of the torsion angles indicated the different flexibilities of their loop struc-tures. The oxirane C6 atom of CA074 was covalently bonded to the Cys29 S^γ atom (C3-S^γ=1.81 Å), where the S-configuration was transformed to the R-form. Concerning the oxirane carbon atom that participates in the covalent bonding with the Cys residue, an acceptable rule has been proposed. The substrate specificities at the Sn (n=1-3) and Sn' (n=1 and 2) subsites of CB, together with the interaction features as to CA074, have been discussed in comparison with the crystal structure of the papain-CA028 (a CA074-related inhibitor) complex.