Ionic liquids as buffer additives in ionic liquid-polyacrylamide gel electrophoresis separation of mixtures of low and high molecular weight proteins

Abstract

This study aims at investigating methodologies for better separation of proteins using novel hydrophobic ionic liquids (ILs). In this regard, hydrophobic ILs [C n PBr] (n ¼ 4, 6, 8) were synthesized and examined in ionic liquidpolyacrylamide gel electrophoresis (IL-PAGE) as buffer additives for separation of catalase (Cat), transferrin (Tf), bovine serum albumin (BSA), ovalbumin (Ova) and a-lactalbumin (a-Lact). The influence of alkyl chain length of the cation of these ILs and their concentration in running and sample buffers on protein separation was investigated. Separation using ILs as additives was achieved at lower concentrations as compared to standard sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The IL concentrations were 100-fold less in sample buffer and 5-fold less in running buffer as compared to conventional SDS-PAGE. The results demonstrated that ILs additives played a role in improving some protein separation, IL-PAGE provided higher resolution and separation efficiency than SDS-PAGE for Tf and Ova. Fluorescence studies were performed in order to understand protein-IL interactions and were used to determine the appropriate IL for use as a buffer additive in PAGE. When compared with standard SDS-PAGE, no heating of sample buffer was required in IL-PAGE, which revealed that proteins could be efficiently denatured by use of IL, which was later confirmed by use of circular dichroism (CD) studies.

Do Ionic Liquids Exhibit the Required Characteristics to Dissolve, Extract, Stabilize, and Purify Proteins? Past-Present-Future Assessment

“…The myoglobin adsorbs on the surface of BPG forming a stable monolayer. The biosensor developed could directly detect the concentration of oxygen in an aqueous solution with a detection limit of 2.3 × 10 −8 M. Hasan et al 366 exploited pyridinium-based ILs, viz. [C n PyrBr] (n = 4, 8), as buffer additive for high-resolution separation of low and high molecular weight proteins, including catalase, transferrin, BSA, ovalbumin, and α-lactalbumin, in IL-polyacrylamide gel electrophoresis as an alternative to the SDS-PAGE technique.…”

“…Besides giving valuable information on the molecular mechanism of IL-protein interactions, the electrolytic nature of ILs as adjuvants has also been exploited for applications in oxygen sensing and IL-PAGE for the separation of proteins. , Ding et al reported that a direct electrochemical response of myoglobin could be observed for oxygen reduction from basal plane graphite (BPG) when [OHC 2 mim]­[BF 4 ] was used as a supporting electrolyte. The myoglobin adsorbs on the surface of BPG forming a stable monolayer.…”

Protein Sieving with Capillary Nanogel Electrophoresis

“…In gel-based sieving, the analyte migration is impeded . In comparison to electrophoresis in a slab gel, electrophoresis in a gel-filled capillary provides sharper peaks and better resolution of biopolymers in a shorter separation time. , The proteins separated by sieving are driven through the gel with a similar electrophoretic transport without any additive or by including a surfactant ,,,, or an ionic liquid , in the gel to bind to the proteins and cause them to have the same charge-to-size ratio. The proteins pass through the porous gel.…”