title: The pH-dependent membrane association of procathepsin L is mediated by a 9-residue sequence within the propeptide.
abstract: 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.
authors.affiliation: Department of Biochemistry and Biophysics, University of North Carolina, Chapel Hill 27599.
authors.forename: G F
authors.initials: GF
authors.lastname: McIntyre
authors.forename: G D
authors.initials: GD
authors.lastname: Godbold
authors.forename: A H
authors.initials: AH
authors.lastname: Erickson
mesh_terms: D016475:3T3 Cells; D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D050260:Carbohydrate Metabolism; D002241:Carbohydrates / Q000737:chemistry; D056668:Cathepsin L; D002403:Cathepsins / Q000378:metabolism*; D002462:Cell Membrane / Q000201:enzymology; D004792:Enzyme Precursors / Q000378:metabolism*; D006735:Horseradish Peroxidase / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007425:Intracellular Membranes / Q000378:metabolism; D008247:Lysosomes / Q000378:metabolism; D008358:Mannose / Q000032:analysis; D051379:Mice; D008861:Microsomes / Q000378:metabolism; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000378:metabolism; D010455:Peptides / Q000378:metabolism; D017386:Sequence Homology, Amino Acid
chemical_list: D002241:Carbohydrates; D004792:Enzyme Precursors; D010446:Peptide Fragments; D010455:Peptides; D006735:Horseradish Peroxidase; D002403:Cathepsins; C063905:procathepsin L; D056668:Cathepsin L; D008358:Mannose