Background
Apolipoprotein E (ApoE) ε4 genotype is the most prevalent risk factor for late-onset Alzheimer’s Disease (AD). Although ApoE4 differs from its non-pathological ApoE3 isoform only by the C112R mutation, the molecular mechanism of its proteinopathy is unknown.
Methods
Here, we reveal the molecular mechanism of ApoE4 aggregation using a combination of experimental and computational techniques, including X-ray crystallography, site-directed mutagenesis, hydrogen-deuterium mass spectrometry (HDX-MS), static light scattering and molecular dynamics simulations. Treatment of ApoE ε3/ε3 and ε4/ε4 cerebral organoids with tramiprosate was used to compare the effect of tramiprosate on ApoE4 aggregation at the cellular level.
Results
We found that C112R substitution in ApoE4 induces long-distance (> 15 Å) conformational changes leading to the formation of a V-shaped dimeric unit that is geometrically different and more aggregation-prone than the ApoE3 structure. AD drug candidate tramiprosate and its metabolite 3-sulfopropanoic acid induce ApoE3-like conformational behavior in ApoE4 and reduce its aggregation propensity. Analysis of ApoE ε4/ε4 cerebral organoids treated with tramiprosate revealed its effect on cholesteryl esters, the storage products of excess cholesterol.
Conclusions
Our results connect the ApoE4 structure with its aggregation propensity, providing a new druggable target for neurodegeneration and ageing.
Graphic Abstract
Allosteric changes imposed by post-translational modifications regulate and differentiate the functions of proteins with intrinsic disorder regions. HDM2 is a hub protein with a large interactome and with different cellular functions. It is best known for its regulation of the p53 tumour suppressor. Under normal cellular conditions, HDM2 ubiquitinates and degrades p53 by the 26S proteasome but after DNA damage, HDM2 switches from a negative to a positive regulator of p53 by binding to p53 mRNA to promote translation of the p53 mRNA. This change in activity is governed by the ataxia telangiectasia mutated kinase via phosphorylation on serine 395 and is mimicked by the S395D phosphomimetic mutant. Here we have used different approaches to show that this event is accompanied by a specific change in the HDM2 structure that affects the HDM2 interactome, such as the N-termini HDM2–p53 protein–protein interaction. These data will give a better understanding of how HDM2 switches from a negative to a positive regulator of p53 and gain new insights into the control of the HDM2 structure and its interactome under different cellular conditions and help identify interphases as potential targets for new drug developments.
Apolipoprotein 4 (APOE) ϵ4 genotype is the most prevalent risk factor for late-onset Alzheimer's Disease (AD). Although APOE4 differs from its non-pathological APOE3 isoform only by the C112R mutation, the mechanism of its proteinopathy is poorly understood. Here, we combine experimental and computational techniques to uncover a domino-like effect of C112R mutation on APOE4 behavior. We found that C112R substitution in APOE4 induces long-distance (>15 Å) conformational changes leading to the formation of a T-shaped dimeric unit that is geometrically different and more aggregation-prone than the APOE3 structure. AD drug candidate tramiprosate and its metabolite 3-sulfopropanoic acid induce APOE3-like conformational behavior in APOE4 and suppress its aggregation propensity. Analysis of APOE ε4/ε4 cerebral organoids treated with tramiprosate revealed its effect on cholesteryl esters, the storage products of excess cholesterol. Overall, our results connect the APOE4 structure with its aggregation propensity, providing a new druggable target for neurodegeneration and ageing.
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