Characterization of Heparin Binding by a Peptide from Amyloid P Component Using Capillary Electrophoresis, Surface Plasmon Resonance, and Isothermal Titration Calorimetry
Authors
Hernaiz, Maria J.
LeBrun, Laurie A.
Wu, Yi
Sen, Jette W.
Linhardt, Robert J.
Heegaard, Niels H.H.
ORCID
https://orcid.org/0000-0003-2219-5833
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Other Contributors
Issue Date
2002-07-22
Keywords
Biology , Chemistry and chemical biology , Chemical and biological engineering , Biomedical engineering
Degree
Terms of Use
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Full Citation
Characterization of Heparin Binding by a Peptide from Amyloid P Component Using Capillary Electrophoresis, Surface Plasmon Resonance, and Isothermal Titration Calorimetry, M.J. Hernaiz, L.A. LeBrun, Y. Wu, J.W. Sen, R.J. Linhardt, N.H.H. Heegaard, European Journal of Biochemistry, 269, 2860-2867. 2002.
Abstract
Synthetic peptides based on amino-acid residues 27-38 of human serum amyloid P component represent a novel type of heparin binders as they do not contain clusters of basic amino acids or other known features associated with protein or peptide heparin binding. Here, we characterize the binding using capillary electrophoresis (CE), surface plasmon resonance (SPR), and isothermal titration calorimetry (ITC). By CE, heparin-binding activity was readily apparent for both a regular peptide and a slightly N-terminally modified form, while a sequence-scrambled peptide had no measurable binding. Dissociation constants in the 1-15 microm range were estimated, but only a minor part of the binding isotherm was covered by the experiments. SPR measurements using immobilized peptides verified heparin binding, the range of the binding constants, and the reduced binding of the sequence-scrambled peptide. Structurally defined heparin oligosaccharides were used to establish that while the tetrasaccharide is too small to exhibit strong binding, little difference in binding strength is observed between hexa- and tetradeca-saccharides. These experiments also confirmed the almost complete lack of activity of the sequence-scrambled peptide. The amino-acid sequence-dependent binding and the importance of a disulfide bond in the peptide were verified by ITC, but the experimental conditions had to be modified because of peptide precipitation and ITC yielded significantly weaker binding constants than the other methods. While the precise function of the peptide in the intact protein remains unclear, the results confirm the specificity of the glycosaminoglycan interaction with regard to peptide sequence by applying two additional biophysical techniques and showing that the N-terminal part of the peptide may be modified without changing the heparin binding capabilities.
Description
European Journal of Biochemistry, 269, 2860-2867
Note : if this item contains full text it may be a preprint, author manuscript, or a Gold OA copy that permits redistribution with a license such as CC BY. The final version is available through the publisher’s platform.
Note : if this item contains full text it may be a preprint, author manuscript, or a Gold OA copy that permits redistribution with a license such as CC BY. The final version is available through the publisher’s platform.
Department
The Linhardt Research Labs.
The Shirley Ann Jackson, Ph.D. Center for Biotechnology and Interdisciplinary Studies (CBIS)
The Shirley Ann Jackson, Ph.D. Center for Biotechnology and Interdisciplinary Studies (CBIS)
Publisher
Relationships
The Linhardt Research Labs Online Collection
Rensselaer Polytechnic Institute, Troy, NY
European Journal of Biochemistry
https://harc.rpi.edu/
Rensselaer Polytechnic Institute, Troy, NY
European Journal of Biochemistry
https://harc.rpi.edu/