CoChat
Sign Up
Cited by 67Alzheimer's disease research and treatmentsProtein Structure and DynamicsProtein Interaction Studies and Fluorescence Analysis

The Association−Dissociation Behavior of the ApoE Proteins: Kinetic and Equilibrium Studies

Open access
Garai, Kanchan, Frieden, Carl·2010·Biochemistry
DOI 10.1021/bi101407mPMC PMC3038193
Open this paper in CoChat — free account

Abstract

The apolipoprotein E family consists of three major protein isoforms: apolipoprotein E4 (ApoE4), ApoE3, and ApoE2. The isoforms, which contain 299 residues, differ only by single-amino acid changes, but of the three, only ApoE4 is a risk factor for Alzheimer’s disease. At micromolar concentrations, lipid-free ApoE exists predominantly as tetramers. In more dilute solutions, lower-molecular mass species predominate. Using fluorescence correlation spectroscopy (FCS), intermolecular fluorescence resonance energy transfer (FRET), and sedimentation methods, we found that the association−dissociation reaction of ApoE can be modeled with a monomer−dimer−tetramer process. Equilibrium constants have been determined from the sedimentation data, while the individual rate constants for association and dissociation were determined by measurement of the kinetics of dissociation of ApoE and are in agreement with the equilibrium constants. Dissociation kinetics as measured by intermolecular FRET show two phases reflecting the dissociation of tetramer to dimer and of dimer to monomer, with dissociation from tetramer to dimer being more rapid than the dissociation from dimer to monomer. The rate constants differ for the different ApoE isoforms, showing that the association−dissociation process is isoform specific. Strikingly, the association rate constants are almost 2 orders of magnitude slower than expected for a diffusion-controlled process. Dissociation kinetics were also monitored by tryptophan fluorescence in the presence of acrylamide and the data found to be consistent with the monomer−dimer−tetramer model. The approach combining multiple methods establishes the reaction scheme of ApoE self-association.

The claims and statistics of this paper haven’t been checked yet.

Run the check — free, no account needed. It reads the open-access text and checks the sources it cites.

A few free checks a day · papers already verified open instantly · results are public

Integrity signals

Registry

Crossref · 10.1021/bi101407m

Cites

7 of 7 references verified0 citation errors · 0 not checkable

Statistics

Claims the paper makes

The claims and statistics of this paper haven’t been checked yet.

Run the check — free, no account needed. It reads the open-access text and checks the sources it cites.

A few free checks a day · papers already verified open instantly · results are public

Claims other papers make about this one

Each is checked against this paper’s own text: does it say what the citing paper says it does?

  • ApoE4 had a greater tendency to aggregate and existed primarily as tetramers, whereas the apoE4-Jac variant consisted mostly of dimers even at a high concentration of apoE (30 μM).

    “Importantly, apoE4 was shown to have greater tendency to aggregate and existed primarily as tetramers (28, 29), whereas apoE4-Jac variant consisted mostly dimers even when a high concentration of apoE (30 μM) was used (Fig. S10A).”

    From <i>APOE3</i> -Jacksonville (V236E) variant reduces self-aggregation and risk of dementia (2021) · cites this paper as [28]

    The Discussion (Differences between the isoforms; page unavailable) reports that ApoE4 is predominantly tetrameric above 2.5 μM, supporting the tetramer portion of the cited clause. It finds only small differences in self-association among the isoforms and does not establish a greater tendency for ApoE4 to aggregate relative to ApoE4-Jac. The dimer finding for ApoE4-Jac is attributed to the citing paper’s own Fig. S10A, not to this source. (LLM, conf=0.84)

    Experimental — accuracy not yet measured

  • The NanoBiT signal reflects the non-covalent, C-terminal-domain-mediated self-association that is common to all three isoforms and has been characterized biophysically as a mixture of dimers, tetramers, and higher-order oligomers in equilibrium.

    “The NanoBiT signal instead reflects the non-covalent, C-terminal-domain-mediated self-association that is common to all three isoforms and has been characterized biophysically as a mixture of dimers, tetramers, and higher-order oligomers in equilibrium (14).”

    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 [14]

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

    Experimental — accuracy not yet measured

  • ApoE has a well-documented tendency to self-associate and form oligomers.

    “ApoE presents particular challenges due to its well-documented tendency to self-associate and form oligomers 40,62 .”

    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 [62]

Self-check

Percentages, sums and reported differences re-computed from the paper’s own numbers.

The self-check needs the paper’s full text.

Cites

Flagged first, then by citations

Research works printed but not linked: 41

  1. [1]Corder EH Saunders AM Strittmatter WJ Schmechel DE Gaskell PC Small GW Roses AD Haines JL Pericak-Vance MA Gene dose of apolipoprotein E type 4 allele and the risk of Alzheimer’s disease in late onset families Science 1993 261 921 923 8346443
  2. [3]Farrer LA Cupples LA Haines JL Hyman B Kukull WA Mayeux R Myers RH Pericak-Vance MA Risch N van Duijn CM Effects of age, sex, and ethnicity on the association between apolipoprotein E genotype and Alzheimer disease. A meta-analysis. APOE and Alzheimer Disease Meta Analysis Consortium Jama 1997 278 1349 1356 9343467
  3. [4]Alberts MJ Graffagnino C McClenny C DeLong D Strittmatter W Saunders AM Roses AD ApoE genotype and survival from intracerebral haemorrhage Lancet 1995 346 575 7658797
  4. [8]Dergunov AD Hoy A Smirnova EA Visvikis S Siest G Charge-based heterogeneity of human plasma lipoproteins at hypertriglyceridemia: capillary isotachophoresis study Int J Biochem Cell Biol 2003 35 530 543 12565713
  5. [9]Chou CY Lin YL Huang YC Sheu SY Lin TH Tsay HJ Chang GG Shiao MS Structural variation in human apolipoprotein E3 and E4: secondary structure, tertiary structure, and size distribution Biophys J 2005 88 455 466 15475580 PMC1305022
  6. [10]Perugini MA Schuck P Howlett GJ Self-association of human apolipoprotein E3 and E4 in the presence and absence of phospholipid J Biol Chem 2000 275 36758 36765 10970893
  7. [12]Patel AB Khumsupan P Narayanaswami V Pyrene fluorescence analysis offers new insights into the conformation of the lipoprotein-binding domain of human apolipoprotein E Biochemistry 2010 49 1766 1775 20073510
  8. [13]Fan D Li Q Korando L Jerome WG Wang J A monomeric human apolipoprotein E carboxyl-terminal domain Biochemistry 2004 43 5055 5064 15109264
  9. [15]Yokoyama S Kawai Y Tajima S Yamamoto A Behavior of human apolipoprotein E in aqueous solutions and at interfaces J Biol Chem 1985 260 16375 16382 4066713
  10. [16]Barbier A Clement-Collin V Dergunov AD Visvikis A Siest G Aggerbeck LP The structure of human apolipoprotein E2, E3 and E4 in solution 1. Tertiary and quaternary structure Biophys Chem 2006 119 158 169 16139946
  11. [17]Munoz F Valles MA Donoso J Echevarria G Garcia Blanco F Kinetic and thermodynamic study of the tetramerization equilibrium of phosphorylase b J Biochem 1983 94 1649 1659 6418736
  12. [18]Kitano H Maeda Y Okubo T Kinetic study of the effects of solvation on the dimerization process of alpha-chymotrypsin Biophys Chem 1989 33 47 54 2720089
  13. [19]Lu HS Chang WC Mendiaz EA Mann MB Langley KE Hsu YR Spontaneous dissociation-association of monomers of the human-stem-cell-factor dimer Biochem J 1995 305 Pt 2 563 568 7530446 PMC1136399
  14. [20]Moore JM Patapoff TW Cromwell ME Kinetics and thermodynamics of dimer formation and dissociation for a recombinant humanized monoclonal antibody to vascular endothelial growth factor Biochemistry 1999 38 13960 13967 10529242
  15. [21]Veronese PK Stafford RP Lucius AL The Escherichia coli ClpA molecular chaperone self-assembles into tetramers Biochemistry 2009 48 9221 9233 19650643
  16. [22]Venezia CF Meany BJ Braz VA Barkley MD Kinetics of association and dissociation of HIV-1 reverse transcriptase subunits Biochemistry 2009 48 9084 9093 19715314 PMC2770954
  17. [23]Hoggett JG Kellett GL Kinetics of the monomer-dimer reaction of yeast hexokinase PI Biochem J 1992 287 Pt 2 567 572 1445216 PMC1133202
  18. [24]Darke PL Jordan SP Hall DL Zugay JA Shafer JA Kuo LC Dissociation and association of the HIV-1 protease dimer subunits: equilibria and rates Biochemistry 1994 33 98 105 8286367
  19. [25]Chen MJ Mayo KH Human platelet factor 4 subunit association/dissociation thermodynamics and kinetics Biochemistry 1991 30 6402 6411 2054346
  20. [26]Garai K Mustafi SM Baban B Frieden C Structural differences between apolipoprotein E3 and E4 as measured by (19)F NMR Protein Sci 2010 19 66 74 19904741 PMC2817840
  21. [27]Magde D Elson EL Webb WW Fluorescence correlation spectroscopy. II. An experimental realization Biopolymers 1974 13 29 61 4818131
  22. [28]Maiti S Haupts U Webb WW Fluorescence correlation spectroscopy: diagnostics for sparse molecules Proc Natl Acad Sci U S A 1997 94 11753 11757 9342306 PMC33774
  23. [29]Garai K Muralidhar M Maiti S Fiber-optic fluorescence correlation spectrometer Appl Opt 2006 45 7538 7542 16983444
  24. [30]Nitsche JM Chang HC Weber PA Nicholson BJ A transient diffusion model yields unitary gap junctional permeabilities from images of cell-to-cell fluorescent dye transfer between Xenopus oocytes Biophys J 2004 86 2058 2077 15041648 PMC1304059
  25. [31]Schuck P Perugini MA Gonzales NR Howlett GJ Schubert D Size-distribution analysis of proteins by analytical ultracentrifugation: strategies and application to model systems Biophys J 2002 82 1096 1111 11806949 PMC1301916
  26. [32]Schuck P Size-distribution analysis of macromolecules by sedimentation velocity ultracentrifugation and lamm equation modeling Biophys J 2000 78 1606 1619 10692345 PMC1300758
  27. [33]Selvin PR Ha T Single-molecule techniques: a laboratory manual 2008 Cold Spring Harbor Laboratory Press Cold Spring Harbor, N.Y.
  28. [34]Johnson KA Simpson ZB Blom T Global kinetic explorer: a new computer program for dynamic simulation and fitting of kinetic data Anal Biochem 2009 387 20 29 19154726
  29. [35]Johnson KA Simpson ZB Blom T FitSpace explorer: an algorithm to evaluate multidimensional parameter space in fitting kinetic data Anal Biochem 2009 387 30 41 19168024
  30. [37]Kellett GL Gutfreund H Reactions of haemoglobin dimers after ligand dissociation Nature 1970 227 921 926 5448994
  31. [38]Pollard TD Cooper JA Actin and actin-binding proteins. A critical evaluation of mechanisms and functions Annu Rev Biochem 1986 55 987 1035 3527055
  32. [39]Noble RW Reichlin M Gibson QH The reactions of antibodies with hemeprotein antgens. The measurement of reaction kinetics and stoichiometry by fluorescence quenching J Biol Chem 1969 244 2403 2411 4977444
  33. [40]Ward ES Gussow D Griffiths AD Jones PT Winter G Binding activities of a repertoire of single immunoglobulin variable domains secreted from Escherichia coli Nature 1989 341 544 546 2677748
  34. [41]Northrup SH Erickson HP Kinetics of protein-protein association explained by Brownian dynamics computer simulation Proc Natl Acad Sci U S A 1992 89 3338 3342 1565624 PMC48862
  35. [42]Hesse C Larsson H Fredman P Minthon L Andreasen N Davidsson P Blennow K Measurement of apolipoprotein E (apoE) in cerebrospinal fluid Neurochem Res 2000 25 511 517 10823584
  36. [43]Schuck P Sedimentation analysis of noninteracting and self-associating solutes using numerical solutions to the Lamm equation Biophys J 1998 75 1503 1512 9726952 PMC1299825
  37. [44]Karbassi F Quiros V Pancholi V Kornblatt MJ Dissociation of the octameric enolase from S. pyogenes--one interface stabilizes another PLoS One 2010 5 e8810 20098674 PMC2809091
  38. [45]Perugini MA Schuck P Howlett GJ Differences in the binding capacity of human apolipoprotein E3 and E4 to size-fractionated lipid emulsions Eur J Biochem 2002 269 5939 5949 12444983
  39. [46]Salter JD Krucinska J Raina J Smith HC Wedekind JE A hydrodynamic analysis of APOBEC3G reveals a monomer-dimer-tetramer self-association that has implications for anti-HIV function Biochemistry 2009 48 10685 10687 19839647 PMC2783380
  40. [47]Brown PH Balbo A Schuck P On the analysis of sedimentation velocity in the study of protein complexes Eur Biophys J 2009 38 1079 1099 19644686 PMC2755746
  41. [48]Brookes E Cao W Demeler B A two-dimensional spectrum analysis for sedimentation velocity experiments of mixtures with heterogeneity in molecular weight and shape Eur Biophys J 2010 39 405 414 19247646 PMC12146834

Cited by

Sorted by citations

Authors
Garai, K., & Frieden, C.
Affiliations
Washington University in St. Louis (US)
Corresponding author
Frieden, Carl
ORCID
Venue
Biochemistry
Publisher
American Chemical Society (ACS)
Published
2010-10-14
Volume
49
Issue
44
Pages
9533-9541
ISSN
  • 0006-2960 (ISSN-L)
  • 1520-4995
DOI
10.1021/bi101407m
PMC
PMC3038193
OpenAlex
W2035467780
Cited by
67
References
7
Citations by year
2026: 6 · 2025: 3 · 2024: 2 · 2023: 4 · 2021: 4 · 2020: 5 · 2019: 5 · 2018: 2 · 2017: 4 · 2016: 3 · 2015: 2 · 2014: 6 · 2013: 7 · 2012: 5
FWCI
4.27
Type
journal article
Language
en
Open access
Locations
  • Biochemistry (publishedVersion)
  • PubMed (publishedVersion)
  • Europe PMC (PubMed Central) (acceptedVersion)
  • PubMed Central (acceptedVersion)
Funders
National Institute of Diabetes and Digestive and Kidney Diseases (DK 13332, R01DK013332)
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 › Protein Interaction Studies and Fluorescence Analysis
Keywords
apolipoprotein E, APOE isoforms, protein self-association, association kinetics, dissociation kinetics, equilibrium constants, Alzheimer's disease risk, tryptophan fluorescence, acrylamide quenching
MeSH terms
Alzheimer Disease, Apolipoproteins E, Humans, Kinetics, Protein Conformation, Spectrometry, Fluorescence, Protein Isoforms, Fluorescence Resonance Energy Transfer, Apolipoprotein E3, Apolipoprotein E4, Apolipoprotein E2, Protein Multimerization

About these checks

Registry, retraction, Cites and statistics results are computed, not judged, with one exception: a printed reference that no index lists is read once by a model before it counts as a citation error. Claims are extracted by a model and kept only when their quote appears verbatim in the text; each cited source is then checked, with a model’s help, for whether it says what the claim relies on. Those verdicts can be wrong — read the claim’s quote and the cited source before relying on one.

Statistics and arithmetic are recomputed, not judged.

Confidence bands are shown only for fields calibrated on our annotated test set. None is calibrated yet, so no band is shown.

What counts as a claim ›How we measure confidence ›How we check this ›

Are you an author, or do you see a problem with this page? Request a review