Affinity-based reverse micellar extraction and separation (ARMES): a facile technique for the purification of peroxidase from soybean hulls.
A new technique for the purification of proteins has been developed which combines the high selectivity of affinity interaction with the scalability and ease of operation of liquid-liquid extraction. The approach is called affinity-based reverse micellar extraction and separation (ARMES). The salient features of ARMES include the following: (1) intraphasic interaction between the ligand and ligate which provides for high ligand utilization; (2) no chemical modification of the ligand is needed; and (3) ease of operation and inherent scalability due to the use of liquid-liquid extraction. This technique has been used to purify the peroxidase from soybean hulls using the lectin concanavalin A (con A) as a sugar-binding affinity ligand. A purification factor of 30 is achieved to provide a nearly pure peroxidase solution (as determined by HPLC and SDS-PAGE) with nearly complete regeneration of the con A ligand. We propose that ARMES will be useful in the facile purification of complex biomolecules such as glycoform protein variants using lectins as affinity ligands and proteins of therapeutic importance using antibodies as affinity ligands.
Paradkar
V M
VM
Department of Chemical and Biochemical Engineering, University of Iowa, Iowa City 52242.
Dordick
J S
JS
D001709:Biotechnology / Q000379:methods; D002623:Chemistry Techniques, Analytical / Q000379:methods; D002846:Chromatography, Affinity / Q000379:methods; D003208:Concanavalin A; D008823:Micelles; D010544:Peroxidases / Q000302:isolation & purification*; D037121:Plant Lectins; D014674:Plant Proteins, Dietary / Q000302:isolation & purification*; D030262:Soybean Proteins; D013025:Glycine max / Q000201:enzymology*
D008823:Micelles; D037121:Plant Lectins; D014674:Plant Proteins, Dietary; D030262:Soybean Proteins; D003208:Concanavalin A; D010544:Peroxidases