Glycosaminoglycan-protein interaction studies using fluorescence spectroscopy

Boothello RS, Al-Horani RA, Desai UR. Glycosaminoglycan-protein interaction studies using fluorescence spectroscopy. Methods Mol Biol. 2015;1229:335-53. doi: 10.1007/978-1-4939-1714-3_27. PMID: 25325964

Abstract
Fluorescence spectroscopy is a quantitative analytical tool that has been extensively used to provide structural and dynamical information on GAG-protein complexes. It possesses major advantages including high sensitivity, relative ease of applicability, and wide range of available fluorescence labels and probes. It has been applied to practically every protein-GAG system through the use of either intrinsic (e.g., Trp) or extrinsic (e.g., a non-covalent fluorophore) probe. For studies involving GAGs, it forms the basis for measurement of dissociation constant of complexes and the stoichiometry of binding, which helps elucidate many other thermodynamic and/or mechanistic parameters. We describe the step-by-step procedure to measure the affinity of GAG-protein complexes, parse the ionic and nonionic components of the free energy of binding, and identify the site of GAG binding through competitive binding experiments.