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Cited by 91Cholesterol and Lipid MetabolismLipoproteins and Cardiovascular HealthLipid Membrane Structure and Behavior

Two-step Mechanism of Binding of Apolipoprotein E to Heparin

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Futamura, Miho, Dhanasekaran, Padmaja, Handa, Tetsurou, Phillips, Michael C., Lund-Katz, Sissel, Saito, Hiroyuki·2005·Journal of Biological Chemistry
DOI 10.1074/jbc.m411719200
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Abstract

The interaction of apolipoprotein E (apoE) with cell-surface heparan sulfate proteoglycans is an important step in the uptake of lipoprotein remnants by the liver. ApoE interacts predominantly with heparin through the N-terminal binding site spanning the residues around 136-150. In this work, surface plasmon resonance analysis was employed to investigate how amphipathic alpha-helix properties and basic residue organization in this region modulate binding of apoE to heparin. The apoE/heparin interaction involves a two-step process; apoE initially binds to heparin with fast association and dissociation rates, followed by a step exhibiting much slower kinetics. Circular dichroism and surface plasmon resonance experiments using a disulfide-linked mutant, in which opening of the N-terminal helix bundle was prevented, demonstrated that there is no major secondary or tertiary structural change in apoE upon heparin binding. Mutations of Lys-146, a key residue for the heparin interaction, greatly reduced the favorable free energy of binding of the first step without affecting the second step, suggesting that electrostatic interaction is involved in the first binding step. Although lipid-free apoE2 tended to bind less than apoE3 and apoE4, there were no significant differences in rate and equilibrium constants of binding among the apoE isoforms in the lipidated state. Discoidal apoE3-phospholipid complexes using a substitution mutant (K143R/K146R) showed similar binding affinity to wild type apoE3, indicating that basic residue specificity is not required for the effective binding of apoE to heparin, unlike its binding to the low density lipoprotein receptor. In addition, disruption of the alpha-helix structure in the apoE heparin binding region led to an increased favorable free energy of binding in the second step, suggesting that hydrophobic interactions contribute to the second binding step. Based on these results, it seems that cell-surface heparan sulfate proteoglycan localizes apoE-enriched remnant lipoproteins to the vicinity of receptors by fast association and dissociation.

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Crossref · 10.1074/jbc.m411719200

Cites

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  • The receptor-binding sites (AA136-150) for LDLR, LRP1, and HSPGs are located in a region conserved between mouse and human APOE.

    “However, the receptor-binding sites (AA136-150) for lipoprotein receptors such as the low-density lipoprotein receptor (LDLR), lipoprotein-related receptor 1 (LRP1), and heparan sulfate proteoglycans (HSPGs) are located in a region conserved between species (Fig. 1a) [11–14].”

    From APOE Christchurch enhances a disease-associated microglial response to plaque but suppresses response to tau pathology (2025) · cites this paper as [13]

    The paper’s introductory discussion (page unavailable) identifies residues 136–150 as the LDLR-binding region and says the same domain is involved in LRP and HSPG binding. It does not state that this region is conserved between mouse and human APOE; searches for mouse–human conservation found no such passage. (LLM, conf=0.88)

    Experimental — accuracy not yet measured

  • The split-luciferase assay relied on the established toxicity of Aβ oligomeric intermediates.

    “based on the known toxicity of the oligomeric intermediates of Aβ35.”

    From Resistance to autosomal dominant Alzheimer’s disease in an APOE3 Christchurch homozygote: a case report (2019) · cites this paper as [35]

    The Experimental Procedures and SPR Measurements sections describe apoE binding to heparin, not a split-luciferase assay. Searches for Aβ oligomers, their toxicity, and split luciferase found no passage in our copy addressing the attributed claim; page numbers were unavailable. (LLM, conf=0.76)

    Experimental — accuracy not yet measured

  • Although both heparin binding sites are functional in separated fragments, only the N-terminal site is available for interaction in both lipid-free and lipidated states of intact apoE.

    “Although both sites are functional in the separated fragments, only the N-terminal site is available for interaction in both the lipid-free and lipidated states of the intact apoE molecule (27, 28).”

    From Role of the N- and C-Terminal Domains in Binding of Apolipoprotein E Isoforms to Heparan Sulfate and Dermatan Sulfate: A Surface Plasmon Resonance Study (2008) · cites this paper as [28]

  • The binding of apoE to heparin is a two-step process; the initial binding involves fast electrostatic interaction, followed by a slower hydrophobic interaction.

    “Using surface plasmon resonance, we previously showed that the binding of apoE to heparin is a two-step process; the initial binding involves fast electrostatic interaction, followed by a slower hydrophobic interaction.”

    From Role of the N- and C-Terminal Domains in Binding of Apolipoprotein E Isoforms to Heparan Sulfate and Dermatan Sulfate: A Surface Plasmon Resonance Study (2008) · cites this paper as [28]

  • A discoidal complex of apoE 22-kDa fragments with DMPC exhibited much higher affinity in binding to heparin compared to the lipid-free form, mainly due to the increased free energy of binding in the first step.

    “Although lipid-free apoE can bind effectively to GAG, we demonstrated previously that a discoidal complex of apoE 22-kDa fragments with DMPC exhibited much higher affinity in binding to heparin compared to the lipid-free form, mainly due to the increased free energy of binding in the first step (28).”

    From Role of the N- and C-Terminal Domains in Binding of Apolipoprotein E Isoforms to Heparan Sulfate and Dermatan Sulfate: A Surface Plasmon Resonance Study (2008) · cites this paper as [28]

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Cites

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Research works printed but not linked: 2

  1. Strittmatter, W. J., and Roses, A. D. (1995) Proc. Natl. Acad. Sci. U. S. A. 92, 4725-4727
  2. Pitas, R. E., Innerarity, T. L., Arnold, K. S., and Mahley, R. W. (1979) Proc. Natl. Acad. Sci. U. S. A. 76, 2311-2315

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Authors
Futamura, M., Dhanasekaran, P., Handa, T., Phillips, M. C., Lund-Katz, S., & Saito, H.
Affiliations
  • Children's Hospital of Philadelphia (US)
  • Kyoto University (JP)
ORCID
Venue
Journal of Biological Chemistry
Publisher
Elsevier BV
Published
2005-02
Volume
280
Issue
7
Pages
5414-5422
ISSN
0021-9258 (ISSN-L)
DOI
10.1074/jbc.m411719200
OpenAlex
W2090843520
Cited by
91
References
56
Citations by year
2026: 2 · 2025: 8 · 2024: 5 · 2023: 4 · 2022: 1 · 2021: 5 · 2020: 5 · 2019: 3 · 2018: 4 · 2017: 5 · 2016: 9 · 2015: 1 · 2014: 7 · 2013: 5 · 2012: 3
FWCI
3.47
Type
journal article
Language
en
Open access
Licence
Locations
  • Journal of Biological Chemistry (publishedVersion, cc-by)
  • PubMed (publishedVersion)
Funders
  • National Institutes of Health
  • Japan Society for the Promotion of Science (12470489, 14572045)
  • National Heart, Lung, and Blood Institute (R01HL056083, HL-56083)
Topics
  • Health Sciences › Medicine › Surgery › Cholesterol and Lipid Metabolism
  • Health Sciences › Medicine › Surgery › Lipoproteins and Cardiovascular Health
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Lipid Membrane Structure and Behavior
Keywords
apolipoprotein E, heparin binding, heparan sulfate proteoglycans, two-step binding mechanism, N-terminal binding site, binding kinetics, electrostatic interactions, hydrophobic interactions, basic residues, APOE isoforms, liver, LDL receptor
MeSH terms
Apolipoproteins E, Binding Sites, Electron Spin Resonance Spectroscopy, Gels, Heparin, Humans, Kinetics, Lysine, Models, Biological, Mutation, Peptide Fragments, Protein Conformation, Sepharose, Thermodynamics, Heparan Sulfate Proteoglycans, Protein Isoforms, Protein Transport, Apolipoprotein E3

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