Effect of protease inhibitors of actinomycetes on lysosomal peptide-hydrolases from swine liver.

small pieces, homogenized with two volumes of 50mMTris buffer (pH 7.4) containing 0.25 Msucrose and 1 mMEDTA, and centrifuged at 700g for 10minutes. After removal of the precipitate (nuclear-debris), the supernatant was further centrifuged at 7,000g for 60minutes. To the resulting sediment (lysosomal-mitochondrial pellet) was added 200ml of cold acetone, and the suspension was stirred for an hour at 4°C. After filtration of the acetone suspension, the filter cake was washed with cold acetone and dried in vacuo. The acetone powder thus obtained (yield 3.2g) was used as a commonsource of cathepsins A, B and D. Before use, an appropriate amount of the acetone powder was extracted with five volumes of 0.1m citrate, pH 5.0. The extract was dialyzed twice (first, 4-hour