2024
DOI: 10.1101/2024.04.03.588023
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Equilibrium dialysis with HPLC detection to measure substrate binding affinity of a non-heme iron halogenase

Elizabeth R. Smithwick,
Ambika Bhagi-Damodaran,
Anoop Rama Damodaran

Abstract: Determination of substrate binding affinity (Kd) is critical to understanding enzyme function. An extensive number of methods have been developed and employed to study ligand/substrate binding, but the best approach depends greatly on the substrate and the enzyme in question. Below we describe how to measure theKdof BesD, a non-heme iron halogenase, for its native substrate lysine using equilibrium dialysis with subsequent detection with High Performance Liquid Chromatography (HPLC). This method can be perform… Show more

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