Activity and stability of α-chymotrypsin in biocompatible ionic liquids: enzyme refolding by triethyl ammonium acetate

Abstract

In view the of wide scope of structural information of biomolecules in biocompatible ionic liquids (ILs) in various applications including chemical and biochemical, it is essential to study the productive preferential interactions between biological macromolecules and biocompatible ILs. We have therefore explored the stability and activity of α-chymotrypsin (CT) in the presence of five ILs from different families, such as triethyl ammonium acetate (TEAA), triethyl ammonium phosphate (TEAP) from ammonium salts, 1-benzyl-3-methylimidazolium chloride ([Bzmim][Cl]), 1-benzyl-3-methylimidazolium tetrafluoroborate ([Bzmim][BF(4)]) from imidazolium salts and tetra-butyl phosphonium bromide (TBPBr) from phosphonium families. Circular dichroism (CD) and UV-vis spectrophotometer experiments were used to study CT stabilization by ILs, related to the associated structural changes and enzyme activity studies, respectively. We observed that all ILs have a dominant contribution to the stabilization of CT. The stability and activity of CT depends on the structural arrangement of the ions of ILs. Our experimental results explicitly elucidate that more hydrophobic imidazolium and phosphonium cations carrying longer alkyl chains of ILs ([Bzmim][Cl], [Bzmim][BF(4)] and TBPBr) were weak stabilizers for CT, while small alkyl chain molecules of triethyl ammonium salts (TEAA and TEAP) are strong stabilizers and therefore more biocompatible for CT stability. Our CD and NMR measurements reveal that TEAA is a refolding additive for CT from a quenched thermal unfolded enzyme structure.

Temperature dependence measurements and molecular interactions for ammonium ionic liquid with N-methyl-2-pyrrolidone

“…ILs are not only a new class of materials, but also they do have a set of physical properties that has sparked increased attention in recent years. The significant attention that in the past decade the scientific community has shown towards mainly due to the great versatility in cation-anion combination and favourable unique properties such as high ionic mobility, non-flammability, and high potential for recycling and extremely low vapour pressure [4][5][6][7]. For these reasons, they rapidly gained interest as greener replacements for traditional volatile organic solvents (VOCs).…”

Ammonium based ionic liquids act as compatible solvents for glycine peptides

“…The potential utilization and applications of ILs are rapidly increasing in all modern scientific fields by several researchers such as separations [13] reactions [14][15][16][17], homogeneous two-phase catalysis [18] and extractions [19]. The activity, structure and stability of protein in aqueous IL solutions have been reported by various researchers [20][21][22][23][24][25].…”

Self-buffering and biocompatible ionic liquid based biological media for enzymatic research

“…61,62 Basically, a-CT is a serine protease enzyme, widely distributed in nature, and known to perform diverse functions. 63 It is widely used to understand the folding/ unfolding mechanism of protein in coslvents. 64 …”