Investigating the effects of Choline-based Deep Eutectic Solvent on bioluminescent reaction of wild type and Arg-rich mutant luciferase

Deep eutectic solvents (DES) are significantly important in enzymatic reactions. These solvents increase the efficient contact between substrate and enzyme and therefore, may increase enzyme stability and activity. Bioluminescence is the production and emission of light by a living organism. The principal chemical reaction in bioluminescence involves the light-emitting pigment luciferin and the enzyme luciferase. The enzyme catalyzes the oxidation of luciferin, which requires the energy-carrying molecule adenosine triphosphate (ATP). Because of its easily detectable product, luciferase has found a wide range of application in biotechnology and molecular biology. However, its low thermostability, low turnover number and high Km for ATP restrict further use of luciferase based systems in commercial application. Newly developed method which can be used to increase the stability of proteins and biocatalysts is to take advantage of Deep eutectic solvents (DESs). One group of Deep eutectic solvents are generally composed of organic salts with hydrogen bond donors, as a result of which hydrogen bonds are formed. In this study, we investigated the effects of these solvents on kinetic properties of wild type and E354R/Arg356 mutant Lampyris turkestanicus luciferases in the presence of choline chloride: glycerol with molar ratio of 2:1 as DES. For this, both wild type and mutant, expressed in Escherichia coli BL21, the protein of interest purified through affinity chromatography and used for kinetic studies. Based on the obtained results, DES has positive effect on the thermostability of wild type and mutant luciferases.