Proteolytic specificity and physiological role of mammalian cathepsin E

点击翻译按钮获取中文摘要

1. A 16.8-kb genomic clone containing the mouse cathepsin E gene was isolated from a 129/SvJ XFix II library. The gene was determined to be 13,8 kb in length and to consist of 9 exons. Highly conservative sequences were identified in the 5'-flnaking region containing potential binding motifs for GATA-transcriptional factors. The mouse cathepsin E gene was assigned to chromosome 1 band F by fluorescence in situ hybridization.2. Molecular cloning of a cDNA for a pepsin inhibitor in the round worm, Ascaris suum, was achieved. The ORF was found to encode a 20-residue potential signal peptide and a 149-residue inhibitor moiety. To obtain the active inhibitor, we constracted a yeast expression vector, pYES2API, containing the inducible galactosidase promoter and a DNA fragment containing a fusion protein of the yeast alpha-factor leader and the Ascaris pepsin inhibitor. The active inhibitor was secreted in the culture medium, the yield being approximately 3 mg/I/day, and purified by a two-step procedure that included HPLC.