Improvement of versatile peroxidase activity and stability by a cholinium-based ionic liquid

Abstract

Deep eutectic solvents (DESs) have been introduced as green solvents and less expensive alternatives for ionic liquids. In the current study, effects of a cholinium based DES, Choline chloride:Glycerol (ChCl:G), on function and structure of versatile peroxidase (VP) were studied. Functional analysis of VP in the presence of various concentrations of ChCl:G revealed lower Km values at lower concentrations and improved activity in neutral pH. According to our results, VP presented higher exposure of hydrophobic patches and formed protein assemblies as revealed by extrinsic fluorescence, fluorescence lifetime, and anisotropy measurements. Circular dichroism and UV–vis spectroscopies demonstrated secondary structural changes and some rearrangements in the heme region. Moreover, ChCl:G seemed to protect VP from thermal denaturation probably by forming a protective layer around VP molecules. In contrast to experimental studies on hydrated ChCl:G, molecular dynamic (MD) simulations of VP in a box of pure ChCl:G suggested minor conformational changes. However, MD studies confirmed the rearrangements of the heme region in the presence of ChCl:G compared to water. According to the results, the authors concluded that lower concentration of ChCl:G could improve VP's activity and kinetic due to inducing some conformational changes, particularly in the heme vicinity, and also by stabilizing the intermediate complex during catalysis.