CRYSTAL-STRUCTURE OF GLYCYL ENDOPEPTIDASE FROM CARICA-PAPAYA - A CYSTEINE ENDOPEPTIDASE OF UNUSUAL SUBSTRATE-SPECIFICITY

Glycyl endopeptidase is a cysteine endopeptidase of the papain family, characterized by specificity for cleavage C-terminal to glycyl residu es only and by resistance to inhibition by members of the cystatin fam ily of cysteine proteinase, inhibitors. Glycyl endopeptidase has been crystallized from high salt with a substrate-like inhibitor covalently bound to the catalytic Cys 25. The structure has been solved by molec ular replacement with the structure of papain and refined at 2.1 Angst rom to an R factor of 0.196 (R(free) = 0.258) with good geometry. The structure of the S-1 substrate binding site of glycyl endopeptidase di ffers from that of papain by the substitution of glycines at residues 23 and 65 in papain, with glutamic acid and arginine, respectively, in glycyl endopeptidase, The side chains of these residues form a barrie r across the binding pocket, effectively excluding substrate residues with large side chains from the S-1 subsite. The constriction of this subsite in glycyl endopeptidase explains the unique specificity of thi s enzyme for cleavage after glycyl residues and is a major component o f its resistance to inhibition by cystatins.