Concentration-Related Response Potentiometric Titrations To Study the Interaction of Small Molecules with Large Biomolecules

In the present paper, the utility of a special potentiometric titration approach for recognition and calculation of biomolecule/small-molecule interactions is reported. This approach is fast, sensitive, reproducible, and inexpensive in comparison to the other methods for the determination of the association constant values (K-a) and the interaction energies (Delta G). The potentiometric titration measurement is based on the use of a classical polymeric membrane indicator electrode in a solution of the small-molecule ligand. The biomolecule is used as a titrant. The potential is measured versus a reference electrode and transformed into a concentration-related signal over the entire concentration interval, also at low concentrations, where the millivolt (y-axis) versus log c(analyte) (x-axis) potentiometric calibration curve is not linear. In the procedure, K-a is calculated for the interaction of cocaine with a cocaine binding aptamer and with an anticocaine antibody. To study the selectivity and cross-reactivity, other oligonucleotides and aptamers are tested, as well as other small ligand molecules such as tetrakis(4-chlorophenyl)borate, metergoline, lidocaine, and bromhexine. The calculated K-a compared favorably to the value reported in the literature using surface plasmon resonance. The potentiometric titration approach called concentration-related response potentiometry is used to study molecular interaction for seven macromolecular target molecules and four small-molecule ligands.