a monomeric variant of insulin degrading enzyme (ide) loses its regulatory properties单体的变体的胰岛素降解酶(ide)失去监管的属性.pdfVIP

A Monomeric Variant of Insulin Degrading Enzyme (IDE) Loses Its Regulatory Properties Eun Suk Song, David W. Rodgers, Louis B. Hersh* Department of Molecular and Cellular Biochemistry and the Center for Structural Biology, University of Kentucky, Lexington, Kentucky, United States of America Abstract Background: Insulin degrading enzyme (IDE) is a key enzyme in the metabolism of both insulin and amyloid beta peptides. IDE is unique in that it is subject to allosteric activation which is hypothesized to occur through an oligomeric structuture. Methodology/Principal Findings: IDE is known to exist as an equilibrium mixture of monomers, dimers, and higher oligomers, with the dimer being the predominant form. Based on the crystal structure of IDE we deleted the putative dimer interface in the C-terminal region, which resulted in a monomeric variant. Monomeric IDE retained enzymatic activity, however instead of the allosteric behavior seen with wild type enzyme it displayed Michaelis-Menten kinetic behavior. With the substrate Abz-GGFLRKHGQ-EDDnp, monomeric IDE retained ,25% of the wild type activity. In contrast with the larger peptide substrates b-endorphin and amyloid b peptide 1–40, monomeric IDE retained only 1 to 0.25% of wild type activity. Unlike wild type IDE neither bradykinin nor dynorphin B-9 activated the monomeric variant of the enzyme. Similarly, monomeric IDE was not activated by polyphosphates under conditions in which the activity of wild type enzyme was increased more than 50 fold. Conclusions/Significance: These findings serve to establish the dimer interface in IDE and demonstrate the requirement for an oligomeric form of the enzyme for its regulatory properties. The data support a mechanism where the binding of activators to oligomeric IDE induces a conformational change that