Haloxyfop was reported to exhibit inhibition effect targeting Mycobacterium tuberculosis and pathogenic parasites. To pave its way for drug development, more research is required to determine the affinities interacting with biological receptors in vivo. In this work, the interactions of Haloxyfop with two model transport proteins were investigated by spectroscopic techniques and theoretical simulation. The interaction mechanism, thermodynamic properties and the impact of Haloxyfop-induced conformational change in serum albumins were revealed by series of fluorescence, UV-Vis absorption and circular dichroic spectroscopy. The specific binding sites were determined by site-competitive replacement experiment. Molecular docking and dynamic simulation provided a visual screening in the microscopic binding mode. The structure of Haloxyfop was roughly divided into three parts that exhibit different covalent interaction affinities. The two isomers of Haloxyfop showed a certain degree of affinity difference. Hydrophobic, polar interaction and π-effect were analyzed in detail, and the surface electrostatic potential energy maps were simulated to provide references. The free energy, calculated by the molecular mechanics-generalized born surface area (MM-GBSA) and molecular mechanics-Poisson Boltzmann surface area (MM-PBSA) methods, was decomposed to per-residues, which intuitively revealed relevant contributions in binding process. The role of water existence was explored through molecular dynamic refinement, and the frontier molecular orbital analysis explored the ionic interaction mechanism in electronic level. In general, multiple chemistry method was adopted to fully unravel the properties of Haloxyfop-binding for the sake of rationalizing the applicability as a therapeutic agent. Communicated by Ramaswamy H. Sarma.
Keywords: Haloxyfop; free energy calculation; molecular simulation; spectroscopy; transport protein.