title: A neglected modulator of insulin-degrading enzyme activity and conformation: The pH.
abstract: Insulin-degrading enzyme (IDE), a ubiquitously expressed zinc metalloprotease, has multiple activities in addition to insulin degradation and its malfunction is believed to connect type 2 diabetes with Alzheimer's disease. IDE has been found in many different cellular compartments, where it may experience significant physio-pathological pH variations. However, the exact role of pH variations on the interplay between enzyme conformations, stability, oligomerization state and catalysis is not understood. Here, we use ESI mass spectrometry, atomic force microscopy, surface plasmon resonance and circular dichroism to investigate the structure-activity relationship of IDE at different pH values. We show that acidic pH affects the ability of the enzyme to bind the substrate and decrease the stability of the protein by inducing an α-helical bundle conformation with a concomitant dissociation of multi-subunit IDE assemblies into monomeric units and loss of activity. These effects suggest a major role played by electrostatic forces in regulating multi-subunit enzyme assembly and function. Our results clearly indicate a pH dependent coupling among enzyme conformation, assembly and stability and suggest that cellular acidosis can have a large effect on IDE oligomerization state, inducing an enzyme inactivation and an altered insulin degradation that could have an impact on insulin signaling.
authors.affiliation: Dipartimento di Scienze Chimiche, Università degli Studi di Catania, Viale Andrea Doria 6, 95125 Catania, Italy. Electronic address: grassog@unict.it.
authors.forename: Giuseppe
authors.initials: G
authors.lastname: Grasso
authors.affiliation: Dipartimento di Scienze Chimiche, Università degli Studi di Catania, Viale Andrea Doria 6, 95125 Catania, Italy.
authors.forename: Cristina
authors.initials: C
authors.lastname: Satriano
authors.affiliation: Istituto Biostrutture e Bioimmagini, CNR, Via P. Gaifami 18, 95126 Catania, Italy.
authors.forename: Danilo
authors.initials: D
authors.lastname: Milardi
mesh_terms: D004789:Enzyme Activation; D006863:Hydrogen-Ion Concentration; D007339:Insulysin / Q000737:chemistry* / Q000378:metabolism*; D008958:Models, Molecular; D011487:Protein Conformation; D013329:Structure-Activity Relationship; D013696:Temperature
chemical_list: D007339:Insulysin
keywords: Acidity; Activity; Enzyme; IDE; Mass spectrometry; Surface plasmon resonance