Subcellular Distribution of Human Leukocytic Cathepsins

1. Methods are presented for the disruption of human peripheral leukocytes by sonication and the separation of the released particulate matter by differential centrifugation.

2. The distribution of catheptic activity in the various fractions was investigated both on material obtained from mixed populations of leukocytes and on that from isolated lymphocytes and granulocytes.

3. All of the leukocytic fractions tested contained cathepsins active at pH 3.5 and pH 8.5.

4. At pH 3.5, the catheptic activity was rather evenly distributed among nuclear (I), and light (IV) and heavy (II) granular fractions, with practically no activity in the soluble fraction (V).

5. At pH 8.5, there was always more catheptic activity in the heavier (II) than the lighter (IV) granules, variable amounts in the nuclear fraction (I) and very little in the supernatant (V).

6. At pH 3.5, the lymphocytic granules showed considerably higher proteolytic specific activity than those from granulocytes.

7. At pH 8.5, all fractions except the heavy granules (II) had higher proteolytic specific activities in granulocytes than in lymphocytes.

8. At pH 3.5, and to a lesser extent at pH 8.5, the light (IV) and heavy (II) granules from chronic lymphatic leukemia cells contained much lower cathepic activity than those from normal lymphocytes.

9. Cathepsins in each subcellular fraction are very labile, losing as much as 80 per cent of their activity upon storage at 4 C. overnight in distilled water.