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Cited by 97Mass Spectrometry Techniques and ApplicationsAlzheimer's disease research and treatmentsAdvanced Proteomics Techniques and Applications

Hydrogen/deuterium exchange and electron-transfer dissociation mass spectrometry determine the interface and dynamics of apolipoprotein E oligomerization

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Huang, Richard Y-C., Garai, Kanchan, Frieden, Carl, Gross, Michael L.·2011·Biochemistry
DOI 10.1021/bi2010027PMC PMC3202646
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Abstract

Apolipoprotein E, a 34 kDa protein, plays a key role in triglyceride and cholesterol metabolism. Of the three common isoforms (ApoE2, -3, and -4), only ApoE4 is a risk factor for Alzheimer's disease. All three isoforms of wild-type ApoE self-associate to form oligomers, a process that may have functional consequences. Although the C-terminal domain, residues 216-299, of ApoE is believed to mediate self-association, the specific residues involved in this process are not known. Here we report the use of hydrogen/deuterium exchange (H/DX) coupled with enzymatic digestion to identify those regions in the sequence of full-length apoE involved in oligomerization. For this determination, we compared the results of H/DX of the wild-type proteins and those of monomeric forms obtained by modifying four residues in the C-terminal domain. The three wild-type and mutant isoforms show similar structures based on their similar H/DX kinetics and extents of exchange. Regions of the C-terminus (residues 230-270) of the ApoE isoforms show significant differences of deuterium uptake between oligomeric and monomeric forms, confirming that oligomerization occurs at these regions. To achieve single amino acid resolution, we examined the extents of H/DX by using electron transfer dissociation (ETD) fragmentation of peptides representing selected regions of both the monomeric and the oligomeric forms of ApoE4. From these experiments, we could identify the specific residues involved in ApoE oligomerization. In addition, our results verify that ApoE4 is composed of a compact structure at its N-terminal domain. Regions of C-terminal domain, however, appear to lack defined structure.

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  • R136S participates in interactions with E231, a residue suggested to participate in the oligomerization of apoE3.

    “it participates in interactions with E231 a residue suggested to participate in the oligomerization of apoE3 based on recent studies [69], [70].”

    From Biophysical Analysis of Apolipoprotein E3 Variants Linked with Development of Type III Hyperlipoproteinemia (2011) · cites this paper as [70]

    The Results and Discussion section identifies a C-terminal region involved in apoE self-association, and the paper reports similar oligomerization-related behavior across the isoforms. It does not identify E231 individually as an oligomerization residue or describe an interaction between R136S and E231. No page number is available in our copy. (LLM, conf=0.78)

    Experimental — accuracy not yet measured

  • R251G is located in the C-terminal domain at a position predicted to lie within the oligomerization interface.

    “R251G is located in the C-terminal domain at a position predicted to lie within the oligomerization interface (12), and the Arg-to-Gly substitution likely directly disrupts the intermolecular contacts that stabilize ApoE4 self-association.”

    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 [12]

    The Results and Discussion identifies the C-terminal domain as important for ApoE self-association, and residue 251 falls within that domain. It does not identify residue 251 as part of the oligomerization interface; the structure discussion says deuterium-uptake information is missing for residues 249–262. The cited paper therefore supports the domain-level location, but not the residue-specific interface prediction. (LLM, conf=0.86)

    Experimental — accuracy not yet measured

  • Val236 is located in the C-terminal amphipathic helix that forms the core of the intermolecular coiled-coil interface.

    “Val236 is located in the C-terminal amphipathic helix that forms the core of the intermolecular coiled-coil interface (10,12), and the introduction of a charged glutamate residue at this position likely disrupts hydrophobic packing.”

    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 [12]

    In Results and Discussion, section a (page unavailable), peptide 230–243, which includes Val236, shows a change associated with ApoE self-association. Section c describes amphipathic helices in the C-terminal domain, but its listed possible helix at 238–266 does not include Val236. The paper therefore supports Val236’s location in a self-association-associated region, not the specific claim that it lies in a helix forming the core of a coiled-coil interface. (LLM, conf=0.84)

    Experimental — accuracy not yet measured

  • The introduction of a charged glutamate residue at position 236 likely disrupts hydrophobic packing.

    “Val236 is located in the C-terminal amphipathic helix that forms the core of the intermolecular coiled-coil interface (10,12), and the introduction of a charged glutamate residue at this position likely disrupts hydrophobic packing.”

    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 [12]

    The Results and Discussion (section a; page unavailable) reports that replacing bulky hydrophobic residues in the C-terminal region with smaller hydrophobic or polar/charged residues can prevent oligomerization. That supports the proposed mechanism in general, but the paper does not examine a Val236-to-glutamate substitution or establish that it disrupts hydrophobic packing at that position. The citations in the printed sentence appear immediately after the preceding description of the interface, rather than after this mechanistic inference. (LLM, conf=0.82)

    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 [12]

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Authors
Huang, R. Y.-C., Garai, K., Frieden, C., & Gross, M. L.
Affiliations
Washington University in St. Louis (US)
Corresponding author
Gross, Michael L.
ORCID
Venue
Biochemistry
Publisher
American Chemical Society (ACS)
Published
2011-10-05
Volume
50
Issue
43
Pages
9273-9282
ISSN
  • 0006-2960 (ISSN-L)
  • 1520-4995
DOI
10.1021/bi2010027
PMC
PMC3202646
OpenAlex
W2069118296
Cited by
97
References
8
Citations by year
2026: 3 · 2025: 3 · 2024: 3 · 2023: 5 · 2022: 4 · 2020: 6 · 2019: 6 · 2018: 8 · 2017: 11 · 2016: 9 · 2015: 7 · 2014: 11 · 2013: 9 · 2012: 9
FWCI
4.25
Type
journal article
Language
en
Open access
Locations
  • Biochemistry (publishedVersion)
  • PubMed (publishedVersion)
  • PubMed Central (acceptedVersion)
Funders
  • National Institutes of Health (5p41rr000954-34)
  • National Institute of General Medical Sciences (P41GM103422)
  • National Center for Research Resources (P41-RR000954, 2P41 RR 000954)
Topics
  • Physical Sciences › Chemistry › Spectroscopy › Mass Spectrometry Techniques and Applications
  • Health Sciences › Medicine › Physiology › Alzheimer's disease research and treatments
  • Physical Sciences › Chemistry › Spectroscopy › Advanced Proteomics Techniques and Applications
Keywords
apolipoprotein E, protein oligomerization, C-terminal domain, APOE ε4, APOE isoforms, hydrogen-deuterium exchange, electron transfer dissociation, protein-protein interaction, Alzheimer's disease risk, protein structure, N-terminal domain
MeSH terms
Alzheimer Disease, Amino Acid Sequence, Apolipoproteins E, Humans, Models, Molecular, Spectrometry, Mass, Electrospray Ionization, Deuterium Exchange Measurement, Mutant Proteins, Apolipoprotein E4, Protein Multimerization

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