Deciphering the conformational change and inhibition of Thermus thermophilus laccase in ionic liquid

Ionic Liquids (ILs)-assisted laccase catalysis is an emerging sustainable technique for biodegradation of aromatic recalcitrant. However, a defined ILs-laccase catalysis is under development, as ILs negatively influence laccase activity. Currently, deciphering laccase inhibition mechanism in ILs has been crucial to improve ILs-laccase based biodegradation. Thus, in-depth molecular analysis of hyperthermophilic Thermus thermophilus (TtL) laccase was studied via molecular dynamics simulation at low (2%) and high (10%) concentrations of ethyl-3-methylimidazolium acetate (EmimOAc). In EmimOAc, TtL’s copper-coordinating sites and loops underwent significant fluctuation. Specifically, Protein Structure Network (PSN) based on cliques/communities showed notable remodeling of TtL’s structural conformation upon acetate (OAc-) binding. At lower EmimOAc concentration, OAc− accumulation at TtL’s active site rigidified the catalytic core and thus encountered competitive inhibition; the high radial distribution of the OAc− observed in 2% EmimOAc also supported this hypothesis. Whereas, at higher EmimOAc concentration, the TtL experienced OAc− induced conformational alteration at noncompetitive binding sites, thus causes maximum loss of TtL’s activity. These speculated OAc− binding sites can be a potential mutational target for better TtL’ s activity in EmimOAc. Overall, the molecular details of TtL’s inhibition in EmimOAc were unraveled, thus enabling engineering of extremophilic laccase for high temperature industrial application.