Non-suppressible insulin-like activity of human serum I. Physicochemical properties, extraction and partial purification

Abstract

1. 1. Human serum insulin-like activity (ILA) measured by the rat adipose tissue assay is only partially neutralized by anti-insulin serum. Since the portionot suppressed by anti-insulin serum (non-suppressible ILA) represents the major part of serum ILA, it seemed of importance to investigate its physiochemical properties, 2. 2. The molecular weight of non-suppressible ILA in serum as estimated by gel filtration was between 70 000 and 150 000. On paper electrophoresis it migrated with the α 2- and β-globulins. 3. 3. A purified material containing 1775 μU non-suppressible ILA per mg was obtained from human plasma by a modified Cohn fractionation procedure followed by extraction in acidic ethanol and by gel filtration. 4. 4. Paper electrophoresis separated the purified material into two components, a basic protein devoid of non-suppressible ILA and a β-globulin containing all the non-suppressible ILA. 5. 5. No insulin A-chain was found in purified non-suppressible ILA by paper electrophoresis in 20% formic acid. 6. 6. Purified non-suppressible ILA was heat stable (80°). It was inactivated by alkali and by reduced glutathione. 7. 7. Gel filtration revealed that at neutral pH non-suppressible ILA formed aggregates of different size, which in 5M acetic acid dissociated into one uniform species of molecules with a molecular weight between 6000 and 10 000. 8. 8. The identity of the purified non-suppressible ILA with that present in native serum has not yet been proved.