Characterization of cleavage enzymes for sterol regulatory element bindingprotein in hamster liver microsomes

Sterol regulatory element binding proteins (SREBP-1 and SREBP-2) are the ke y transcription factors for the regulation of the cellular cholesterol leve l. To identify proteolytic enzymes for SREBPs, a fluorogenic peptide substr ate, MOCAc-GrRSVLSFK(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-GRS VL-aldehyde with an IC50 of 40 nM. This peptidase separated into three peak s of approximately 400 kDa, 60 kDa, and 30 kDa (Mp400, Mp60 and Mp30 respec tively) upon gel permeation chromatography. Mp30 was purified to apparent h omogeneity with an M-r of 32 kDa. The partial amino acid sequence of Mp30 p ossessed homology to cathepsin B (EC 3.4.22.1). A 109 kDa protein band on S DS-PAGE which corresponded to Mp400 exhibited homology to neprilysin (EC 3. 4.24.11) in partial amino acid sequence. These findings suggest several deg radative pathways for SREBP in liver microsome membranes. (C) 1999 Academic Press.