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Purification and processing of rat liver procathepsin B
T Kawabata1, Y Nishimura, M Higaki
1Division of Physiological Chemistry, Faculty of Pharmaceutical Sciences, Kyushu University, Fukuoka.
Journal of Biochemistry
|March 1, 1993
Summary
This study reveals that cathepsin D plays a crucial role in activating procathepsin B within rat liver lysosomes. This finding is key to understanding lysosomal enzyme processing and function.
Area of Science:
- Biochemistry
- Cell Biology
- Molecular Biology
Background:
- Lysosomal enzymes undergo complex intracellular processing for activation.
- Cathepsin B is a key lysosomal protease involved in various cellular functions.
- Understanding the precise activation mechanism of procathepsin B is essential.
Purpose of the Study:
- To elucidate the specific mechanism of intracellular processing and activation of procathepsin B.
- To identify the enzymes responsible for procathepsin B maturation in rat liver lysosomes.
Main Methods:
- Purification of procathepsin B from rat liver microsomal contents.
- Enzymatic activity assays and immunoblot analysis.
- Treatment of lysosomal contents to generate enzyme-free fractions.
- NH2-terminal sequencing of processed cathepsin B forms.
Main Results:
- Purified procathepsin B (39 kDa) exhibited no enzymatic activity.
- Incubation with cathepsin B-free lysosomal contents activated procathepsin B to a 31 kDa intermediate form.
- Cathepsin D was identified as essential for this processing and activation event.
- Pepstatin inhibited the processing and activation of procathepsin B.
Conclusions:
- Cathepsin D is critically involved in the processing and activation of procathepsin B in rat liver lysosomes.
- The activation involves conversion to a 31 kDa intermediate form, preceding the mature 29 kDa single-chain form.
- This study provides significant insights into the regulation of lysosomal protease activity.