Protein-Flavonoid Interaction Studies by a Taylor Dispersion Surface Plasmon Resonance (SPR) Technique: A Novel Method to Assess Biomolecular Interactions

Vachali PP, Li B, Besch B, Bernstein PS, 6(6), Biosensors, 2016

Flavonoids are a class of plant-derived polyphenolic compounds. Recent studies indicate that flavonoids have many health benefits including possible anti-cancer and anti-inflammatory properties. Human serum albumin (HSA) is a globular protein present in the blood plasma that functions as a primary carrier of many pharmaceutically important compounds. Glutathione S-transferase pi isoform-1 (GSTP1) is an enzyme that plays a key role in detoxification of xenobiotics with reduced glutathione. Described herein is the assessment of the binding interactions of four flavonoids (kaempferol, luteolin, quercetin, and resveratrol) with HAS and GSTP1, using Taylor dispersion surface plasmon resonance (SPR) assay format. All binding assays involved a Pall ForteBio Pioneer SPR instrument. Polycarboxylate hydrogel sensor chip surfaces were immobilized with HAS and GSTP1 using the standard amine-coupling chemistry. Each of the four flavonoids were injected in running buffer using Taylor diffusion gradient injection mode also known as OneStep injection method. As an assay validation approach, the OneStep method was compared with conventional fixed concentration injection (FCI) method using the interaction between HSA and quercetin. Affinity (KD) and kinetic (ka, kd) constants were determined for the protein-flavonoid interactions. Binding affinities obtained from both OneStep method and conventional fixed concentration method are comparable. Overall results of this study suggest that the OneStep method is compatible with both kinetic and equilibrium analyses of biomolecular interactions.

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