Multi-step processing of procathepsin L in vitro.
The proteolytic processes involved in the conversion of procathepsin L to cathepsin L on a negatively charged surface, dextran sulfate, were studied. Upon incubation for 30 min at 37 degrees C, pH 5.5 with dextran-sulfate and dithiothreitol, purified procathepsin L showed maximal activation and, correspondingly, the complete conversion to the 30 kDa, single chain mature form of enzyme was observed. In contrast, incubation under the same conditions on ice rather than at 37 degrees C for 30 or 60 min resulted in partial proteolysis to produce a 31 kDa form without a significant increase in activity. Amino terminal amino acid sequence analyses showed that the 30 kDa form obtained by incubation at 37 degrees C corresponds to the purified form of mature cathepsin L with a 2 amino acid extension at the amino terminal, and that the 31 kDa form generated by incubation on ice possesses a 6 amino acid amino terminal extension, suggesting that the activation and processing of procathepsin L are different processes, and that 4 amino acid residues (Glu-Pro-Leu-Met) at the carboxyterminal in the propeptide function to prevent the activation of processed cathepsin L.
Ishidoh
K
K
Department of Biochemistry, Juntendo University School of Medicine, Tokyo, Japan.
Kominami
E
E
D016475:3T3 Cells; D000595:Amino Acid Sequence; D000818:Animals; D056668:Cathepsin L; D002403:Cathepsins / Q000302:isolation & purification / Q000378:metabolism*; D002461:Cell Line, Transformed; D003546:Cysteine Endopeptidases; D016264:Dextran Sulfate; D004591:Electrophoresis, Polyacrylamide Gel; D010450:Endopeptidases; D004789:Enzyme Activation; D004792:Enzyme Precursors / Q000302:isolation & purification / Q000378:metabolism*; D011905:Genes, ras; D007700:Kinetics; D051379:Mice; D008969:Molecular Sequence Data; D008970:Molecular Weight; D011499:Protein Processing, Post-Translational
D004792:Enzyme Precursors; D016264:Dextran Sulfate; D002403:Cathepsins; D010450:Endopeptidases; D003546:Cysteine Endopeptidases; C063905:procathepsin L; D056668:Cathepsin L; C534278:Ctsl protein, mouse