Mechanistic insights into lysozyme interaction with MWCNTs and cholinium-based ionic liquids: A tripartite IL–NP–protein interaction study supported by molecular dynamics simulations

Abstract

An improved understanding of the cooperative and antagonistic impacts of ionic liquids (ILs) and nanoparticles (NPs) on proteins is necessary for the progress of biocatalysis, biosensing and nanobiotechnology. Here we, for the first time, analyze an important aspect of the mechanism of interaction of multiwalled carbon nanotubes (MWCNTs) and cholinium-based ILs in terms of lysozyme (Lys) conformational stability. A combination of biophysical methods such as UV-vis spectroscopy, fluorescence spectroscopy, circular dichroism (CD) spectroscopy and dynamic light scattering (DLS) with in silico methods (molecular docking and 100-ns molecular dynamics (MD) simulations) was used to explore the structural changes at the level of secondary and tertiary organization of the Lys. In the presence of MWCNTs, Lys gained stability by promoting enhanced ordering of the α-helical structure and restricting the mobility of the some of the aromatic residues, while ILs such as choline chloride ([ChCl]) and choline acetate ([ChAc]) exerted concentration and anion dependent destabilizing effects. Considerable synergistic stabilization was exhibited by the MWCNT–ChCl–Lys system, with [ChCl] losing destabilizing effects in the presence of MWCNTs, leading to more compact and folded structure of the protein. In the MWCNT–ChAc–Lys system, the stabilization was weak, and no protection from acetate ion-induced unfolding was provided due to the acetate anion's chaotropic properties and its role as a strong hydrogen bond acceptor. MD simulations confirmed that MWCNTs have stable binding to Trp-rich regions of Lys through hydrophobic π–π interactions. They showed the cooperative role of [ChCl] in sustaining the hydration and hydrogen-bond networks. These findings grant the first mechanistic insight into the tripartite IL–NP–protein interactions and thus the promise of choline-based IL–nanomaterial combinations as biocompatible stabilizing agents in protein-based technologies.