Effect of Hemolysis on the Concentration of Insulin in Serum Determined by RIA and IRMA

Insulin-degrading enzyme (IDE; EC 3.4.99.45) was first described 40 years ago (1). It is widely distributed in various tissues, including red blood cells (RBC) (2)(3)(4). IDE may not play a key role in insulin metabolism, and fundamental questions on the biological role of IDE remain (2)(3). Recently, IDE was characterized as a peroxisomal protease (3). The specificity of IDE is selective: Only insulin and transforming growth factor-α ( K m ≠ 0.1 μmol/L) are good substrates; insulin-like growth factor 1 and proinsulin are poor substrates (2)(3)(4). On denaturing polyacrylamide gels, IDE appears as a single polypeptide of 110 kDa (4), but in nonreducing conditions, IDE has an M r of 300 000, suggesting that the enzyme exists in polymer form (4). The cleavage sites indicate that IDE recognizes the tertiary structure rather than a particular amino acid sequence (2). Inhibitors of IDE include p -hydroxymercuribenzoate (0.1 mmol/L), p -chloromercuriphenylsulfonic acid (pcMPS, 0.1 mmol/L), bacitracin (1 g/L), N -ethylmaleimide (1 mmol/L), 1,10-phenanthroline (1 mmol/L), EDTA (5 mmol/L), and diamide (5 mmol/L) (4)(5)(6). The degradation of insulin by IDE is not inhibited by lysosomal enzyme inhibitors like aprotinin (500 000 kU/L) or leupeptin (0.1 g/L) (4)(6). Although most insulin …