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Cited by 48Alzheimer's disease research and treatmentsProtein Structure and DynamicsLipid Membrane Structure and Behavior

Apolipoprotein E4 has extensive conformational heterogeneity in lipid-free and lipid-bound forms

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Stuchell-Brereton, Melissa D., Zimmerman, Maxwell I., Miller, Justin J., Mallimadugula, Upasana L., Incicco, J. Jeremías, Roy, Debjit, Smith, Louis G., Cubuk, Jasmine +5 more·2023·Proceedings of the National Academy of Sciences
DOI 10.1073/pnas.2215371120PMC PMC9963066

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Abstract

The ε4-allele variant of apolipoprotein E (ApoE4) is the strongest genetic risk factor for Alzheimer’s disease, although it only differs from its neutral counterpart ApoE3 by a single amino acid substitution. While ApoE4 influences the formation of plaques and neurofibrillary tangles, the structural determinants of pathogenicity remain undetermined due to limited structural information. Previous studies have led to conflicting models of the C-terminal region positioning with respect to the N-terminal domain across isoforms largely because the data are potentially confounded by the presence of heterogeneous oligomers. Here, we apply a combination of single-molecule spectroscopy and molecular dynamics simulations to construct an atomically detailed model of monomeric ApoE4 and probe the effect of lipid association. Importantly, our approach overcomes previous limitations by allowing us to work at picomolar concentrations where only the monomer is present. Our data reveal that ApoE4 is far more disordered and extended than previously thought and retains significant conformational heterogeneity after binding lipids. Comparing the proximity of the N- and C-terminal domains across the three major isoforms (ApoE4, ApoE3, and ApoE2) suggests that all maintain heterogeneous conformations in their monomeric form, with ApoE2 adopting a slightly more compact ensemble. Overall, these data provide a foundation for understanding how ApoE4 differs from nonpathogenic and protective variants of the protein.

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  • Thermal stability may be a particularly sensitive indicator of structural perturbation in the helical bundle and domain-domain interaction regions.

    “suggests that thermal stability may be a particularly sensitive indicator of structural perturbation in these regions 22,24,33,41,51 .”

    From Utilizing a cell culture based novel cellular thermal shift assay to understand the isoform-dependent thermal stability of ApoE variants (2026) · cites this paper as [24]

    The Results section, “Folding and Stability of the Four-Helix Bundle” (page not available), relates measured stability to changes in the helical bundle; the Discussion, “Folding Equilibrium of Lipid-Free ApoE4” (page not available), also describes associated changes in the hinge and terminal regions. But the paper measures unfolding through chemical denaturant titration, not thermal stability, and does not establish thermal stability as a particularly sensitive indicator of perturbation. (LLM, conf=0.84)

    Experimental — accuracy not yet measured

  • The R61T mutant, which is predicted to disrupt domain interaction without altering the C-terminal sequence, normalizes signal in this assay to ApoE3 levels, providing mechanistic evidence that the N-terminal domain interaction of ApoE4 impacts self-association.

    “The observation that the mutant R61T, which is predicted to disrupt domain interaction without altering the C-terminal sequence, normalizes signal in this assay to ApoE3 levels provides mechanistic evidence that while the C-terminal domain of ApoE is undoubtedly crucial for ApoE oligomerization, the N-terminal domain interaction of ApoE4 impacts self-association (11,12,24,25).”

    From APOE Isoform-Dependent Self-Association Measured by a Split-Luciferase Complementation Assay: Differential Effects of Disease-Risk and Protective Variants (2026) · cites this paper as [24]

    Split into 2 atoms — 0 confirmed, 1 partially supported, 0 disputed, 0 misattributed, 1 unverifiable. Expand the claim to see per-atom verdicts.

    Experimental — accuracy not yet measured

  • ApoE lipidation shifts the monomer-oligomer equilibrium towards monomer.

    “ApoE lipidation is known to shift the monomeroligomer equilibrium towards monomer (18,24,35)”

    From APOE Isoform-Dependent Self-Association Measured by a Split-Luciferase Complementation Assay: Differential Effects of Disease-Risk and Protective Variants (2026) · cites this paper as [24]

    The paper reports that monomeric ApoE4 binds lipids and, in Results > Lipid Association of ApoE4 (page not available), finds one protein per lipid-bound state. It does not show that lipidation shifts the monomer–oligomer equilibrium toward monomer; the reported equilibrium is between lipid-bound conformations, not oligomeric states. (LLM, conf=0.78)

    Experimental — accuracy not yet measured

  • Monomeric ApoE4 adopts conformations distinct from those observed in oligomeric forms.

    “recent single-molecule studies have shown that monomeric ApoE4 adopts conformations distinct from those observed in oligomeric forms 24 .”

    From Utilizing a cell culture based novel cellular thermal shift assay to understand the isoform-dependent thermal stability of ApoE variants (2026) · cites this paper as [24]

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Cited by

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Authors
Stuchell-Brereton, M. D., Zimmerman, M. I., Miller, J. J., Mallimadugula, U. L., Incicco, J. J., Roy, D., Smith, L. G., Cubuk, J., Baban, B., DeKoster, G. T., Frieden, C., Bowman, G. R., & Soranno, A.
Affiliations
  • Department of Biochemistry and Molecular Biophysics, Washington University in St. Louis, Saint Louis, MO 63110
  • Washington University in St. Louis (US)
Corresponding author
Soranno, Andrea
ORCID
Venue
Proceedings of the National Academy of Sciences
Publisher
National Academy of Sciences
Published
2023-02-07
Volume
120
Issue
7
Pages
e2215371120-e2215371120
ISSN
  • 0027-8424 (ISSN-L)
  • 1091-6490
DOI
10.1073/pnas.2215371120
PMC
PMC9963066
OpenAlex
W4319333065
Cited by
48
References
9
Citations by year
2026: 10 · 2025: 15 · 2024: 13 · 2023: 10
FWCI
8.52
Type
journal article
Language
en
Open access
Licence
Locations
  • Proceedings of the National Academy of Sciences (publishedVersion, cc-by)
  • bioRxiv (Cold Spring Harbor Laboratory) (acceptedVersion)
  • PubMed Central (publishedVersion, cc-by)
  • PubMed (publishedVersion)
  • Digital Commons@Becker (Washington University School of Medicine) (submittedVersion, cc-by)
Funders
  • HHS | NIH | National Institute on Aging (P30AG066444-35.2, R01AG062837, U19AG069701, RF1AG067194, T32AG05851804)
  • Ruth K. Broad Biomedical Research Foundation (Extramural Award)
  • Alzheimer's Association (AARG-591858)
  • American Federation for Aging Research (RAG18224)
  • BrightFocus Foundation (A2020382S)
  • David and Lucile Packard Foundation (Packard Fellowship for Science and Engineering)
  • National Science Foundation (Career Award MCB-1552471)
Topics
  • Health Sciences › Medicine › Physiology › Alzheimer's disease research and treatments
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Protein Structure and Dynamics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Lipid Membrane Structure and Behavior
Keywords
APOE ε4, apolipoprotein E isoforms, conformational heterogeneity, protein disorder, lipid binding, Alzheimer's disease risk, monomeric protein structure, N-terminal domain, C-terminal domain, protein-lipid interactions, single-molecule spectroscopy, molecular dynamics simulation
MeSH terms
Apolipoproteins E, Protein Conformation, Protein Isoforms, Apolipoprotein E3, Apolipoprotein E4, Apolipoprotein E2

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