Characterization of the interaction between human lactoferrin and lomefloxacin at physiological condition: Multi-spectroscopic and modeling description

The interaction between lomefloxacin (LMF) and human lactoferrin (Hlf) was studied by using fluorescence, circular dichroism (CD) spectroscopic and molecular modeling measurements. By the fluorescence quenching results, it was found that the binding constant K-A=8.69 x 10(5) L mol(-1), and number of binding sites,n=1.75 at physiological condition. Experimental results observed showed that the binding of LMF to Hit induced conformational changes of Hlf. The participation of tyrosyl and tryptophanyl residues of protein was also estimated in the drug-Hlf complex by synchronous fluorescence. The quantitative analysis data of far-UV CD spectra from that of the alpha-helix 37.4% in free Hlf to 30.2% in the LMF-Hlf complex further confirmed that secondary structure of the protein was changed by LMF. Near-UV CD showed perturbations around tryptophan and tyrosine residues which involves perturbations of tertiary structure. The thermodynamic parameters like, Delta H degrees and Delta S degrees, have been calculated to be 63.411 kj mol(-1) and 231.104 j mol(-1) K-1, respectively. Thermodynamic analysis showed that hydrophobic interactions were the main force in the binding site but the hydrogen bonding and electrostatic interaction could not be excluded which in agreement with the result of molecular docking study. The distance r between donor and acceptor was obtained according to fluorescence resonance energy transfer (FRET) and found to be 1.78 nm. The interaction between LMF and Hlf has been verified as consistent with the static quenching procedure and the quenching mechanism is related to the energy transfer. Furthermore, the study of molecular modeling that LMF could bind to the alpha-helixes between Pro145-Asn152 and Phe167-Gln172 regions and hydrophobic interaction was the major acting force for the binding site, which was in agreement with the thermodynamic analysis. (C) 2010 Elsevier B.V. All rights reserved.