Investigation of the interaction of pepsin with ionic liquids by using fluorescence spectroscopy.

The molecular mechanism of the interaction between pepsin and two typical ionic liquids (ILs), 1-butyl-3-methylimidazolium chloride ([C4mim]Cl) and 1-octyl-3-methylimidazolium chloride ([C8mim]Cl), was investigated with fluorescence spectroscopy, ultraviolet absorption, and circular dichroism spectroscopy at a pH value of 1.6. The results suggest that ILs could quench the intrinsic fluorescence of pepsin, probably via a dynamic quenching mechanism. The fluorescence quenching constants were determined by employing the classic Stern-Volmer equation. The constant values are very small, indicating that only a very weak interaction between ILs and pepsin exists. The Gibbs free-energy change, enthalpy change (ΔH), and entropy change (ΔS) during the interaction of pepsin and ILs were estimated. Positive values of ΔH and ΔS indicate that the interaction between ILs and pepsin is mainly driven by hydrophobic interaction. Synchronous and three-dimensional fluorescence spectra demonstrate that the addition of ILs (0-0.20 mol L(-1) for each IL) does not bring apparent changes to the microenvironments of tyrosine and tryptophan residues. Activity experiments show that the activity of pepsin is concentration dependent; higher concentrations of ILs (>0.22 mol L(-1) for [C8mim]Cl and >0.30 mol L(-1) for [C4mim]Cl) cause the remarkable reduction of enzyme activity. The presence of ILs also does not improve the thermal stability of pepsin.

Interaction of an amino-functionalized ionic liquid with enzymes: a fluorescence spectroscopy study.

The interaction of an amino-functionalized ionic liquid, 1-(2-aminoethyl)-3-butylimidazolium bromide ([NH(2)C(2)C(4)im]Br) with two enzymes, pepsin and papain was investigated using fluorescence spectroscopic technique. It is found that [NH(2)C(2)C(4)im]Br has strong ability to quench the intrinsic fluorescence of pepsin and papain. Quenching mechanisms are considered as static quenching for papain and dynamic quenching for pepsin, respectively. The binding constants and the number of binding sites (n) of [NH(2)C(2)C(4)im]Br to papain were calculated at different temperatures. The thermodynamic parameters such as free energy change (ΔG), enthalpy change (ΔH) and entropy change (ΔS), were calculated by thermodynamic equations. The values of ΔG, ΔH and ΔS suggest that interaction of [NH(2)C(2)C(4)im]Br with the two enzymes is spontaneous. Hydrogen bonding and van der Waals interactions play important roles in the binding process of [NH(2)C(2)C(4)im]Br to papain. However, hydrophobic interaction is the main driving force for the interaction of [NH(2)C(2)C(4)im]Br with pepsin. The results of three-dimensional fluorescence spectra show that [NH(2)C(2)C(4)im]Br has no obvious effects on the polypeptide structures of the two enzymes. Additionally, the [NH(2)C(2)C(4)im]Br-containing system can slightly increase the activities of the two enzymes.