CROSS-CLASS INHIBITION OF THE CYSTEINE PROTEINASES CATHEPSIN-K, CATHEPSIN-L, AND CATHEPSIN-S BY THE SERPIN SQUAMOUS-CELL CARCINOMA ANTIGEN-1 - A KINETIC-ANALYSIS

The human squamous cell carcinoma antigens (SCCA) 1 and 2 are tandemly arrayed genes that encode two high-molecular-weight serine proteinase inhibitors (serpins). Although these proteins are 92% identical, diff erences in their reactive site loops suggest that they inhibit differe nt types of proteinases. Our previous studies show that SCCA2 inhibits chymotrypsin-like serine proteinases [Schick et al. (1997) J. Biol. C hem. 272, 1849-1855]. We now show that, unlike SCCA2, SCCA1 lacks inhi bitory activity against any of the more common types of serine protein ases but is a potent cross-class inhibitor of the archetypal lysosomal cysteine proteinases cathepsins K, L, and S. Kinetic analysis reveale d that SCCA1 interacted with cathepsins K, L, and S at 1:1 stoichiomet ry and with second-order rate constants greater than or equal to 1 x 1 0(5) M-1 s(-1). These rate constants were comparable to those obtained with the prototypical physiological cysteine proteinase inhibitor, cy statin C. Also relative to cystatin C, SCCA1 was a more potent inhibit or of cathepsin K-mediated elastolytic activity by forming longer live d inhibitor-proteinase complexes. The t(1/2) of SCCA1-cathepsin S comp lexes was >1155 min, whereas that of cystatin C-cathepsin complexes wa s 55 min. Cleavage between the Gly and Ser residues of the reactive si te loop and detection of a stable SCCA1-cathepsin S complex by sodium dodecyl sulfate-polyacrylamide gel electrophoresis suggested that the serpin interacted with the cysteine proteinase in a manner similar to that observed for typical serpin-serine proteinase interactions. These data suggest that, contingent upon their reactive site loop sequences , mammalian serpins, in general, utilize their dynamic tertiary struct ure to trap proteinases from more than one mechanistic class and that SCCA1, in particular, may be involved in a novel inhibitory pathway ai med at regulating a powerful array of lysosomal cysteine proteinases.