Insulin-degrading enzyme (IDE) is a zinc metalloprotease that selectively degrades biologically important substrates associated with type 2 diabetes and Alzheimer’s disease (AD). As such, IDE is an attractive target for therapeutic innovations. A major requirement is an understanding of how other molecules present in cells regulate the activity of the enzyme toward insulin, IDE’s most important physiologically relevant substrate. Previous kinetic studies of the IDE-dependent degradation of insulin in the presence of potential regulators have used iodinated insulin, a chemical modification that has been shown to alter the biological and biochemical properties of insulin. Here, we present a novel kinetic assay that takes advantage of the loss of helical circular dichroic signals of insulin with IDE-dependent degradation. As proof of concept, the resulting Michaelis–Menten kinetic constants accurately predict the known regulation of IDE by adenosine triphosphate (ATP). Intriguingly, we found that when Mg2+ is present with ATP, the regulation is abolished. The implication of this result for the development of preventative and therapeutic strategies for AD is discussed. We anticipate that the new assay presented here will lead to the identification of other small molecules that regulate the activity of IDE toward insulin.

Alzheimer’s disease (AD), the most common cause of dementia in the elderly, is the sixth leading cause of death in the United States. We hypothesize that the impaired clearance of Aβ42 from the brain is partly responsible for the onset of sporadic AD. In this work, we evaluated the activity of insulin-degrading enzyme (IDE) toward Aβ42 in the presence of resveratrol, a polyphenol found in red wine and grape juice. By liquid chromatography/mass spectrometry, we identified initial cleavage sites in the absence and presence of resveratrol that carry biological relevance connected to the amyloidogenic properties of Aβ42. Incubation with resveratrol results in a substantial increase in Aβ42 fragmentation compared to the control, signifying that the polyphenol sustains IDE-dependent degradation of Aβ42 and its fragments. Our findings suggest that therapeutic and/or preventative approaches combining resveratrol and IDE may hold promise for sporadic AD.

The insulin-degrading enzyme (IDE) possesses a strong ability to degrade insulin and Aβ42 that has been linked to the neurodegeneration in Alzheimer’s disease (AD). Given this, an attractive IDE-centric strategy for the development of therapeutics for AD is to boost IDE’s activity for the clearance of Aβ42 without offsetting insulin proteostasis. Recently, we showed that resveratrol enhances IDE’s activity toward Aβ42. In this work, we used a combination of chromatographic and spectroscopic techniques to investigate the effects of resveratrol on IDE’s activity toward insulin. For comparison, we also studied epigallocatechin-3-gallate (EGCG). Our results show that the two polyphenols affect the IDE-dependent degradation of insulin in different ways: EGCG inhibits IDE while resveratrol has no effect. These findings suggest that polyphenols provide a path for developing therapeutic strategies that can selectively target IDE substrate specificity.

Alzheimer's disease (AD) is the most common neurodegenerative disease resulting in dementia. It is characterized pathologically by extracellular amyloid plaques composed mainly of deposited Aβ42 and intracellular neurofibrillary tangles formed by hyperphosphorylated tau protein. Recent clinical trials targeting Aβ have failed, suggesting that other polypeptides produced from the amyloid-β precursor protein (APP) may be involved in AD. An attractive polypeptide is AICD57, the longest APP intracellular domain (AICD) coproduced with Aβ42. Here, we show that AICD57 forms micelle-like assemblies that are proteolyzed by insulin-degrading enzyme (IDE), indicating that AICD57 monomers are in dynamic equilibrium with AICD57 assemblies. The N-terminal part of AICD57 monomer is not degraded, but its C-terminal part is hydrolyzed, particularly in the YENPTY motif that has been associated with the hyperphosphorylation of tau. Therefore, sustaining IDE activity well into old age holds promise for regulating levels of not only Aβ but also AICD in the aging brain.

No abstract