Abstract
BackgroundInsulin degrading enzyme (IDE) is responsible for the metabolism of insulin and plays a role in clearance of the Aβ peptide associated with Alzheimer's disease. Unlike most proteolytic enzymes, IDE, which consists of four structurally related domains and exists primarily as a dimer, exhibits allosteric kinetics, being activated by both small substrate peptides and polyphosphates such as ATP.Principal FindingsThe crystal structure of a catalytically compromised mutant of IDE has electron density for peptide ligands bound at the active site in domain 1 and a distal site in domain 2. Mutating residues in the distal site eliminates allosteric kinetics and activation by a small peptide, as well as greatly reducing activation by ATP, demonstrating that this site plays a key role in allostery. Comparison of the peptide bound IDE structure (using a low activity E111F IDE mutant) with unliganded wild type IDE shows a change in the interface between two halves of the clamshell-like molecule, which may enhance enzyme activity by altering the equilibrium between closed and open conformations. In addition, changes in the dimer interface suggest a basis for communication between subunits.Conclusions/SignificanceOur findings indicate that a region remote from the active site mediates allosteric activation of insulysin by peptides. Activation may involve a small conformational change that weakens the interface between two halves of the enzyme.
Highlights
Insulysin (EC 3.4.24.56; referred to as insulin-degrading enzyme or Insulin degrading enzyme (IDE)) is a zinc metallopeptidase in the M16 family that has been extensively studied because of its role in cellular insulin degradation [1] and amyloid beta peptide catabolism [2,3]
The importance of IDE in regulating insulin metabolism became evident from studies of the GK rat model of type II diabetes mellitus [4] where variants of IDE with reduced catalytic activity were found to be associated with elevated insulin levels
Overview of the rat IDE (rIDE) Structure The structures of wild type and E111F rIDE are very similar to the published structures of peptide bound and unliganded human IDE [23,24,25,26]
Summary
Insulysin (EC 3.4.24.56; referred to as insulin-degrading enzyme or IDE) is a zinc metallopeptidase in the M16 family that has been extensively studied because of its role in cellular insulin degradation [1] and amyloid beta peptide catabolism [2,3]. IDE exists predominantly as a dimer in equilibrium with tetramers and to a lesser extent monomers [15,16] It is unique among the enzymes comprising the zinc metallopeptidase M16 family in that it exhibits allosteric kinetic behavior with substrate peptides increasing its activity [16,17,18,19,20]. IDE contains a distinct regulatory site first reported by Camberos and coworkers [21] that binds nucleoside triphosphates and alters IDE activity. This site was shown to be distinct from the active site and to primarily interact with the triphosphate moiety [15]. IDE, which consists of four structurally related domains and exists primarily as a dimer, exhibits allosteric kinetics, being activated by both small substrate peptides and polyphosphates such as ATP
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