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Affinity extraction of proteins with a reversed micellar system composed of Cibacron Blue‐modified lecithin
Author(s) -
Sun Yan,
Ichikawa Sosaku,
Sugiura Shinji,
Furusaki Shintaro
Publication year - 1998
Publication title -
biotechnology and bioengineering
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.136
H-Index - 189
eISSN - 1097-0290
pISSN - 0006-3592
DOI - 10.1002/(sici)1097-0290(19980405)58:1<58::aid-bit6>3.0.co;2-t
Subject(s) - lecithin , chromatography , chemistry , extraction (chemistry) , biochemistry
Crude soybean lecithin was used as a novel surfactant to form reversed micelles in n ‐hexane. Cibacron Blue F‐3GA (CB) was directly immobilized to the reversed micelles by a two‐phase reaction. The reversed micellar system without CB showed low solubilizing capacity for low molecular weight proteins, lysozyme, and cytochrome c due to the weak electrostatic interactions. The introduction of CB significantly increased the solubilization of lysozyme because of its affinity binding to CB but showed no effect on the solubilization of cytochrome c since it did not bind to CB. Although bovine serum albumin had an affinity for CB, it was not extracted to the reversed micelles containing CB because its high molecular weight resulted in a significant steric hindrance effect. Thus the reversed micellar system had a high selectivity resulting from both biospecific and steric hindrance effects. The extraction yield of lysozyme decreased significantly with increasing ionic strength. Therefore, the back extraction of lysozyme was carried out using a stripping solution with an ionic strength of 0.865 mol/L. The overall recovery yield of lysozyme after back extraction could be increased to 87% by stripping for 2 h. The recovered lysozyme exhibited an activity equivalent to native lysozyme, and its secondary structure was also unchanged. © 1998 John Wiley & Sons, Inc. Biotechnol Bioeng 58:58–64, 1998.