Interplay of Multiple Interaction Forces: Binding of Norfloxacin to Human Serum Albumin

Abstract

Herein, the binding interaction of a potential chemotherapeutic antibacterial drug norfloxacin (NOF) with a serum transport protein, human serum albumin (HSA), is investigated. The prototropic transformation of the drug (NOF) is found to be remarkably modified following interaction with the protein as manifested through significant modulations of the photophysics of the drug. The predominant zwitterionic form of NOF in aqueous buffer phase undergoes transformation to the cationic form within the protein-encapsulated state. This implies the possible role of electrostatic interaction force in NOF-HSA binding. This postulate is further substantiated from the effect of ionic strength on the interaction process. To this end, the detailed study of the thermodynamics of the drug-protein interaction process from isothermal titration calorimetric (ITC) experiments is found to unfold the signature of electrostatic as well as hydrophobic interaction forces underlying the binding process. Thus, interplay of more than one interaction forces is argued to be responsible for the overall drug-protein binding. The ITC results reveal an important finding in terms of enthalpy-entropy compensation (EEC) characterizing the NOF-HSA binding. The effect of drug-binding on the native protein conformation has also been evaluated from circular dichroism (CD) spectroscopy which unveils partial rupture of the protein secondary structure. In conjunction to this, the functionality of the native protein (in terms of esterase-like activity) is found to be lowered as a result of binding with NOF. The AutoDock-based docking simulation unravels the probable binding location of NOF within the hydrophilic subdomain IA of HSA. The present program also focuses on exploring the dynamical aspects of the drug-protein interaction scenario. The rotational-relaxation dynamics of the protein-bound drug reveals the not-so-common "dip-and-rise" pattern.

Interaction of human serum albumin with uremic toxins: a thermodynamic study

“…Thus, calorimetric and spectroscopic methods have proven to be very suitable to characterized binding thermodynamics of ligands to proteins, namely ITC 10,12,13,[22][23][24][25] and uorescence spectroscopy. 10,12,13,[23][24][25] Other groups characterized binding isotherms by equilibrium dialysis (ED), 3,4,13,26,27 capillary electrophoresis (CE) [28][29][30] and chromatographic techniques.…”

“…10,12,13,[23][24][25] Other groups characterized binding isotherms by equilibrium dialysis (ED), 3,4,13,26,27 capillary electrophoresis (CE) [28][29][30] and chromatographic techniques. 6,27,31 Up to now, however, only a few studies have explored the dependence on temperature.…”

“…6,27,31 Up to now, however, only a few studies have explored the dependence on temperature. 11,13,22,24 Much less is known about the dependence of the binding strength on ionic strength 24,32 in detail. However, Böhringer et al have recently demonstrated in an in vitro study that the fraction of uremic toxins was signi-cantly decreased through hypertonic predilution hemodial-tration.…”

“…† In general, the binding affinity decreases with increasing temperature as has been observed in several cases of adsorption of small molecules to HSA. 3,13,24,27 For example, Zaidi et al have Table 1 Binding free energies DG bi of PhAA binding to native and modified HSA calculated according to eqn (9). i denotes the first and second binding process in the SBS model studied interaction between hippuric acid (HA) and HSA and their dependence on temperature using steady state uores-cence quenching measurements and ITC.…”

Interaction of Indole Derivative with Human Serum Albumin: A Combined Spectroscopic and Molecular Dynamics Study

“…The effectiveness of a drug as a therapeutic agent is measured by its protein binding property. Due to vast significance of this study, investigation on the binding of drugs to carrier proteins (serum proteins) has been an active area of research …”