Interaction between Fleroxacin and Lysozyme by Using Multi-Spectral Techniques and Molecular Docking

The interaction of fleroxacin (FLE) with extracelluar protein (lysozyme, LYSO) was investigated by using multi-spectral techniques and molecular docking in vitro. Fluorescence spectra studies indicated that FLE quenched LYSO fluorescence in a static mode with binding constants (K-a) of 4. 10 X 10(4) and 0. 74 X 104 L . mol(-1) at 298 and 310 K, respectively. The thermodynamic parameters demonstrated that hydrogen bonds and van der Waals forces played the major role in the binding process. Based on the Forster theory of nonradiative energy transfer, the binding distance (r) between FLE and the inner tryptophan residues of LYSO was calculated to be 3. 16 nm. The Hill's coefficient (n(H)) was calculated by using involved equations, implying the negative cooperativity between them. Circular dichroism spectra (CD) indicated the secondary structure of LYSO was partially destroyed by FLE with the alpha-helix percentage decreasing from 21. 1% to 8. 8%. UV-Vis spectral, synchronous fluorescence and three-dimensional fluorescence spectra revealed the binding interaction could cause conformational and micro-environmental changes of LYSO. The results of molecular docking showed that FLE was mainly bound in the active site hinge region where ASP-52, TRP-62 and TRP-63 were located, and well supported the thermodynamic results. Besides, FIE made the activity of lysozyme decrease with the increasing concentration of fleroxacin, indicating that structural changes in lysozyme can cause the inhibition of lysozyme activity. The work clarified the interaction mechanism of FLE with LYSO at molecular level.