TitleAnion activation site of insulin-degrading enzyme.
Publication TypeJournal Article
Year of Publication2012
AuthorsNoinaj N, Song E S, Bhasin S, Alper BJ, Schmidt WK, Hersh LB, Rodgers DW
JournalJ Biol Chem
Date Published2012 Jan 2
KeywordsAdenosine Triphosphate, Animals, Anions, Binding Sites, Cell Line, Crystallography, X-Ray, Enzyme Activation, Insulysin, Intracellular Space, Models, Molecular, Mutagenesis, Mutation, Protein Conformation, Rats
AbstractInsulin-degrading enzyme (IDE) (insulysin) is a zinc metallopeptidase that metabolizes several bioactive peptides, including insulin and the amyloid β peptide. IDE is an unusual metallopeptidase in that it is allosterically activated by both small peptides and anions, such as ATP. Here, we report that the ATP-binding site is located on a portion of the substrate binding chamber wall arising largely from domain 4 of the four-domain IDE. Two variants having residues in this site mutated, IDEK898A,K899A,S901A and IDER429S, both show greatly decreased activation by the polyphosphate anions ATP and PPPi. IDEK898A,K899A,S901A is also deficient in activation by small peptides, suggesting a possible mechanistic link between the two types of allosteric activation. Sodium chloride at high concentrations can also activate IDE. There are no observable differences in average conformation between the IDE-ATP complex and unliganded IDE, but regions of the active site and C-terminal domain do show increased crystallographic thermal factors in the complex, suggesting an effect on dynamics. Activation by ATP is shown to be independent of the ATP hydrolysis activity reported for the enzyme. We also report that IDEK898A,K899A,S901A has reduced intracellular function relative to unmodified IDE, consistent with a possible role for anion activation of IDE activity in vivo. Together, the data suggest a model in which the binding of anions activates by reducing the electrostatic attraction between the two halves of the enzyme, shifting the partitioning between open and closed conformations of IDE toward the open form.
Alternate JournalJ. Biol. Chem.
PubMed ID22049080