Extractive Solubilization, Structural Change, and Functional Conversion of Cytochrome c in Ionic Liquids via Crown Ether Complexation

Abstract

This article reports on the extraction behavior of heme proteins from an aqueous phase into ionic liquids (ILs) with dicyclohexano-18-crown-6 (DCH18C6), and the structure-function relationship of cytochrome c (Cyt-c) dissolved in ILs. We have found that DCH18C6 enables transfer of Lys-rich proteins into ILs via supramolecular complexation. The hydrophobicity and functional groups of ILs have a great influence on protein partitioning, and a hydroxyl group-containing IL with DCH18C6 is capable of the quantitative partitioning of Cyt-c. On the other hand, protein transfer using conventional organic solvents is negligibly small. UV-visible, CD, and resonance Raman spectroscopic characterizations indicate that the sixth ligand Met 80 in the heme group of the Cyt-c-DCH18C6 complex in IL is replaced by other amino acid residues of the peptide chain and that a non-natural, six-coordinate, low-spin ferric heme structure is induced in IL. Solubilization of Cyt-c in IL causes the environmental change of the heme vicinity of Cyt-c, which triggers the functional conversion of Cyt-c from an electron-transfer protein to peroxidase. The Cyt-c-DCH18C6 complex in IL provides remarkably high peroxidase activity compared with native Cyt-c, because of enhancement of the affinity for H2O2.

Mutual Solubility of Hydrophobic Ionic Liquids and Water in Liquid-Liquid Two-phase Systems for Analytical Chemistry

“…[15][16][17][18][19][20][21][22][23][24][25][26][27][28][29][30] Among many possibilities of applying ionic liquids, IL-water (W) two-phase systems that consist of an aqueous or a nonpolar molecular solvent and an ionic liquid are of considerable relevance with analytical chemistry and separation technology. A direct application of IL-W two-phase systems is the extraction of neutral compounds of relatively low molecular weights, [31][32][33][34][35][36][37][38][39][40][41][42][43][44][45][46][47] biopolymers 44,[48][49][50][51] and ions. Several reviews on extraction with ILs are available.…”

Supramolecular Structures in the Presence of Ionic Liquids

“…Compared to the conventional organic solvents, ILs enabled quantitative recovery of amino acids and eliminated emulsion formation, which usually occurs when cationic or anionic extractants are used. Shimojo et al (Shimojo et al, 2006a;Shimojo et al, 2006b) investigated a hydroxyl-groupcontaining IL with DCH18C6 was capable of quantitative partitioning of heme protein cytochrome c (Cyt-c) via supramolecular complexation and that the resulting Cyt-c-DCH18C6 complex acts as a homogeneous biocatalyst in ILs. While using conventional organic solvents the protein transfer was negligibly small.…”

Enhanced Extraction of Amino Compounds Using Dicyclohexyl-18-crown-6 as a Ligand in an Aqueous Two-phase System

“…In a similar manner, 18-crown-6 recognizes also the protonated primary amino groups of a protein, resulting in the formation of a supramolecular complex between the crown ether molecules and the protein [19][20][21][22]. Recently, extraction of a cationic protein cytochrome c into an ionic liquid using a crown ether as the extractant has been accomplished [23,24]. Similarly, we recently found that the extraction behavior of cytochrome c is influenced by adding a crown ether as the ligand in the Li 2 SO 4 (lithium sulfate)/PEG ATPS [25].…”