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Fluorescent dyes as probes to study lipid-binding proteins

Authors

  • Alexander V. Pastukhov,

    1. Department of Biochemistry and Molecular Biology, Pennsylvania State University College of Medicine, Hershey, Pennsylvania
    Current affiliation:
    1. Center 4 Discovery of Drugs and Diagnostics, University of Central Florida, 12722 Research Parkway, Orlando, FL 32826
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  • Ira J. Ropson

    Corresponding author
    1. Department of Biochemistry and Molecular Biology, Pennsylvania State University College of Medicine, Hershey, Pennsylvania
    • Department of Biochemistry and Molecular Biology, Pennsylvania State University College of Medicine, Hershey, PA 17033
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Abstract

We studied the equilibrium binding of two hydrophobic fluorescent dyes, ANS and bisANS, to four members of a family of intracellular lipid-binding proteins: IFABP, CRABP I, CRABP II, and ILBP. The spectral and binding parameters for the probes bound to the proteins were determined. Typically, there was a single binding site on each protein for the ligands. However, IFABP cooperatively bound a second bisANS molecule in the binding pocket. Comparative analysis of affinities and spectral characteristics for the two probes allowed us to examine the contributions of electrostatic and hydrophobic interactions to the binding process, and to address some aspects of the internal structure of the studied proteins. Proteins 2003;53:000–000. © 2003 Wiley-Liss, Inc.

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