The pH-dependent membrane association of procathepsin L is mediated by a 9-residue sequence within the propeptide.

The lysosomal proprotease procathepsin L binds to mouse fibroblast microsomal membranes at pH 5, but mature active cathepsin L does not (McIntyre, G.F., and Erickson, A. H. (1991) J. Biol. Chem. 266, 15438-15445). This binding is not dependent on N-linked carbohydrate as procathepsin L synthesized in cells treated with tunicamycin still shows pH-dependent membrane association. These results suggest that the propeptide (Thr18-Lys113) of the cysteine protease mediates its pH-dependent membrane association. Synthetic peptides containing either 24 or 9 residues from the N-terminal portion of the mouse procathepsin L propeptide inhibited the binding of mouse procathepsin L to microsomal membranes at pH 5. In contrast, the pH-dependent membrane association was not inhibited either by a scrambled version of the 24-residue peptide, in which 3 adjacent residues likely to be positively charged at pH 5 were dispersed, or by a second control peptide containing the 11 N-terminal residues from mature mouse cathepsin L. The 24-residue peptide chemically coupled to horseradish peroxidase bound to microsomes at pH 5, but not at pH 7. On ligand blots, the same conjugate bound specifically to a 43-kDa integral membrane protein, identifying the microsomal protein that mediates the proenzyme binding. The 9-residue propeptide sequence that inhibits the membrane association of procathepsin L at pH 5 resembles the vacuolar sorting sequences in the propeptides of yeast proteinase A and carboxypeptidase Y. This suggests that the membrane association of procathepsin L may play a role in the transport of the proenzyme to lysosomes, the vacuolar equivalent in mammalian cells.