Label-free evaluation of small-molecule-protein interaction using magnetic capture and electrochemical detection

摘要

The evaluation of interaction between small molecules and protein is an important step in the discovery of new drugs and to study complex biological systems. In this work, an alternative method was presented to evaluate small-molecule-protein interaction by using ligand capture by protein-coated magnetic particles (MPs) and disposable electrochemical cells. The interaction study was conducted using 10-gingerol from ginger rhizome and a transmembrane protein alpha V beta 3 integrin. Initially, the electrochemical behavior of the natural compound 10-gingerol was evaluated with the disposable carbon-based electrodes and presented an irreversible oxidation process controlled by diffusion. The analytical curve for 10-gingerol was obtained in the range of 1.0 to 20.0 mu mol L-1, with limit of detection of 0.26 mu mol L-1. Then MPs coated with V3 integrin were incubated with standard solutions and extracts of ginger rhizome for 10-gingerol capture and separation. The bioconjugate obtained was dropped to the disposable electrochemical cells, keeping a permanent magnet behind the working electrode, and the binding process was evaluated by the electrochemical detection of 10-gingerol. The assay method proposed was also employed to calculate the 10-gingerol-alpha V beta 3 integrin association constant, which was calculated as 4.3 x 10(7) M-1. The method proposed proved to be a good label-free alternative to ligand-protein interaction studies.