title: Stabilization of enzymes in ionic liquids via modification of enzyme charge.
abstract: Due to the propensity of ionic liquids (ILs) to inactivate enzymes, the development of strategies to improve enzyme utility in these solvents is critical to fully exploit ILs for biocatalysis. We have developed a strategy to broadly improve enzyme utility in ILs based on elucidating the effect of charge modifications on the function of enzymes in IL environments. Results of stability studies in aqueous-IL mixtures indicated a clear connection between the ratio of enzyme-containing positive-to-negative sites and enzyme stability in ILs. Stability studies of the effect of [BMIM][Cl] and [EMIM][EtSO4 ] on chymotrypsin specifically found an optimum ratio of positively-charged amine-to-negatively-charged acid groups (0.39). At this ratio, the half-life of chymotrypsin was increased 1.6- and 4.3-fold relative to wild-type chymotrypsin in [BMIM][Cl] and [EMIM][EtSO4 ], respectively. The half-lives of lipase and papain were similarly increased as much as 4.0 and 2.4-fold, respectively, in [BMIM][Cl] by modifying the ratio of positive-to-negative sites of each enzyme. More generally, the results of stability studies found that modifications that reduce the ratio of enzyme-containing positive-to-negative sites improve enzyme stability in ILs. Understanding the impact of charge modification on enzyme stability in ILs may ultimately be exploited to rationally engineer enzymes for improved function in IL environments.
authors.affiliation: Department of Chemical and Biological Engineering, University of Colorado, Boulder, CO 80309, USA.
authors.forename: Erik M
authors.initials: EM
authors.lastname: Nordwald
authors.forename: Joel L
authors.initials: JL
authors.lastname: Kaar
mesh_terms: D000588:Amines / Q000737:chemistry; D000818:Animals; D055162:Biocatalysis; D002417:Cattle; D002918:Chymotrypsin / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D004795:Enzyme Stability / Q000235:genetics*; D007093:Imidazoles / Q000737:chemistry; D052578:Ionic Liquids / Q000737:chemistry*; D015202:Protein Engineering / Q000379:methods*
chemical_list: D000588:Amines; D007093:Imidazoles; D052578:Ionic Liquids; D002918:Chymotrypsin
keywords: biocatalysis; enzyme engineering; enzyme modification; ionic liquids