A DIRECT INHIBITORY EFFECT OF INSULIN ON A CYTOSOLIC PROTEOLYTIC COMPLEX CONTAINING INSULIN-DEGRADING ENZYME AND MULTICATALYTIC PROTEINASE

The insulin degrading enzyme (IDE) and the multicatalytic proteinase ( MCP) can be isolated as components of a cytosolic proteolytic complex. IDE is the primary enzyme involved in cellular degradation of insulin , and insulin has been shown to interact with cytosolic IDE. MCP is be lieved to be important in nonubiquitin pathways of cellular protein de gradation. Insulin has a dose- and time-dependent inhibitory effect on MCP degradation of N-succinyl-Leu-Leu-Val-Tyr 7-amido-4-methylcoumari n (LLVY), a substrate for MCP. Proinsulin also inhibits LLVY degradati on in a dose dependent manner. The effect of insulin is immediate as m easured in a continuously monitored assay of LLVY degradation. purific ation of the IDE MCP complex using a variety of approaches, including affinity and conventional chromatography, retains the insulin effect o n LLVY degradation as long as the complex remains intact. After ion-ex change chromatography, which separates IDE and MCP, insulin no longer has an inhibitory effect. Recombination of purified IDE and MCP does n ot restore the effect of insulin, but inclusion of additional componen ts from the ion exchange column does. These results support the existe nce of a functional cytosolic complex that contains IDE and MCP. Insul in interacts with IDE and alters the activity of MCP, suggesting a fun ctional relationship between these two components and a mechanism for an intracellular action of insulin.