Related Experiment Video
Updated: Aug 15, 2026
In vivo and in vitro Studies of Adaptor-clathrin Interaction
Published on: January 26, 2011
Molecular cloning of cDNA for SPase, a monkey cathepsin L orthologue
Toru Nishinaka1, Jun Song, Kristen Lum
1Oral Biology & Medicine, School of Dentistry University of California, Los Angeles, CA, 90095, USA.
Abstract:
SPase is a cysteine protease isolated from an African green monkey kidney cell line, CV-1, and has selective cleavage activity toward transcription factor SP-1 and retinoblastoma susceptibility gene product RB. In this study, a cDNA encoding SPase was cloned from a cDNA library prepared from CV-1 cells. The cDNA clone encodes 333 amino acids and is 96.5% identical to human cathepsin L at the nucleotide and amino acid sequence levels. SPase appears to be translated as a preproenzyme based on the comparison between the deduced amino acid sequence and the N-terminal sequence of the purified enzyme. Northern blot analysis exhibited the considerably higher expression of SPase in CV-1 cells compared with COS-1 cells, showing a good correlation with enzymatic activity in these cell lines. Bacterially expressed SPase protein exhibited proteolytic activity toward SP-1 and RB proteins. These observations suggest that SPase is a monkey cathepsin L orthologue.
Insights
SPase, a monkey cysteine protease, cleaves transcription factor SP-1 and retinoblastoma protein RB. This study identifies SPase as a monkey cathepsin L orthologue, crucial for understanding primate protease functions.
Area of Science:
- Biochemistry
- Molecular Biology
- Genetics
Background:
- SPase is a cysteine protease from African green monkey kidney cells (CV-1) with selective cleavage activity.
- It targets transcription factor SP-1 and the retinoblastoma susceptibility gene product (RB).
Purpose of the Study:
- To clone the cDNA encoding SPase from CV-1 cells.
- To characterize SPase and determine its relationship to human cathepsin L.
Main Methods:
- cDNA library screening and cloning.
- Sequence analysis and comparison with human cathepsin L.
- Northern blot analysis for gene expression.
- Bacterial expression and proteolytic activity assays.
Main Results:
- A cDNA encoding 333 amino acids was cloned, showing 96.5% identity to human cathepsin L.
- SPase is likely translated as a preproenzyme.
- SPase expression is significantly higher in CV-1 cells than COS-1 cells, correlating with enzymatic activity.
- Bacterially expressed SPase demonstrated proteolytic activity against SP-1 and RB.
Conclusions:
- SPase is identified as a monkey cathepsin L orthologue.
- The findings provide insights into primate protease function and regulation.
- SPase's selective cleavage activity has implications for cellular processes involving SP-1 and RB.
