Counterpointing Scenarioson the Fate of Different Prototropic Formsof Norfloxacin Housed in the Pocket of Lysozyme: The NonelectrostaticInteractions in the Protein Interior Are in the Controlling Role onthe Prototropic Equilibria of the Guest

摘要

Herein, we report a comprehensive study on the interaction of three protomeric forms of the antibacterial drug norfloxacin (nfx) with the enzymatic protein human lysozyme (lyz). Norfloxacin, having the option for two-stage acid–base equilibria, converts from cationic (nfx⁺) to zwitterionic (nfx^±) form, followed by an anionic (nfx^–) species, with increasing pH. Among these protomeric forms, lysozyme binds nfx^± most robustly, whereas nfx^– has a weak association and nfx⁺ does not show any interaction. In lysozyme, the location of the drug was ascertained by competitive binding assay with 8-anilino-1-naphthalenesulfonate, and this was further examined with molecular docking simulation. The binding process was found to be primarily governed by hydrogen bonding and van der Waals interactions. The study has further revealed that preferential binding of nfx^± by the protein over nfx^– led to a switchover of nfx^– to nfx^±; and the resulting increased population of nfx^± over the other is beneficial for the pharmacological activity of the drug in terms of its accumulation in the target bacterial cells. The present study accomplishes two important objectives. It holdssignificance regarding the differential interaction of multiprotomericdrugs with biomolecules, such as proteins, enzymes, lipid membranes,etc., and also on such biomolecule-assisted alteration of the acid–baseequilibrium and consequent bioavailability of the drug. The findingsare useful from the viewpoints of dispensation, distribution, andmetabolism of any prototropic drug in living systems as they encounterseveral biomolecules in vivo. Another importance of this work stemsfrom the study of comparative binding responses of lysozyme towarda drug existing in multiple forms depending on its protonation statesor some other chemical processes.