Characterization of Cleavage Enzymes for Sterol Regulatory Element Binding Protein in Hamster Liver Microsomes

• Published in: Biochemical and biophysical research communications Vol. 258; no. 3; pp. 542 - 547
• Main Authors: Yamaguchi, Mineko, Shirai, Heigoro, Sato, Ryuichiro, Kawabe, Yoshiki, Fukuda, Rie, Kodama, Tatsuhiko, Hamakubo, Takao
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 19.05.1999
• Subjects: Amino Acid Sequence; Animals; cathepsin B; CCAAT-Enhancer-Binding Proteins; cholesterol; Cricetinae; DNA-Binding Proteins - metabolism; endoplasmic reticulum; Humans; Male; Mesocricetus; Microsomes, Liver - enzymology; Molecular Sequence Data; neprilysin; Nuclear Proteins - metabolism; Peptide Hydrolases - chemistry; Peptide Hydrolases - metabolism; protease; Protease Inhibitors - pharmacology; SREBP; Sterol Regulatory Element Binding Protein 1; Transcription Factors; protease; cholesterol; SREBP; neprilysin; cathepsin B; endoplasmic reticulum
• ISSN: 0006-291X; 1090-2104
• DOI: 10.1006/bbrc.1999.0679

Sterol regulatory element binding proteins (SREBP-1 and SREBP-2) are the key transcription factors for the regulation of the cellular cholesterol level.To identify proteolytic enzymes for SREBPs, a fluorogenic peptide substrate, MOCAc-GRSVLSFK(Dnp)rr-NH2, was synthesized according to the proposed cleavage site of human SREBP-2. In microsome fractions from hamster liver, we found a peptidase activity inhibitable by the synthetic inhibitor Ac-GRSVL-aldehyde with an IC50of 40 nM. This peptidase separated into three peaks of approximately 400 kDa, 60 kDa, and 30 kDa (Mp400, Mp60 and Mp30 respectively) upon gel permeation chromatography. Mp30 was purified to apparent homogeneity with anMrof 32 kDa. The partial amino acid sequence of Mp30 possessed homology to cathepsin B (EC 3.4.22.1). A 109 kDa protein band on SDS-PAGE which corresponded to Mp400 exhibited homology to neprilysin (EC 3.4.24.11) in partial amino acid sequence. These findings suggest several degradative pathways for SREBP in liver microsome membranes.