Applicability of the stoichiometric displacement model to description of the retention behavior of charged-fusion proteins during fast protein liquid chromatography.
The applicability of the stoichiometric displacement model (SDM) to description of the retention behavior of charged-fusion proteins in large ion exchange resin (approximately 90 microm diameter) packed column was studied. Proteins were characterized by SDM for isocratic elution. The parameters were subsequently used to evaluate their suitability in predicting protein retention and peak width under gradient elution. The proteins were beta-glucuronidase (GUS) and its fusions with polypeptides of 5, 10 and 15 aspartic acids at the C-terminal of the wild-type GUS. Predictions of retention time were within 10% of the experiment results. The plate number obtained at high salt concentration from isocratic elution was used as a first estimate for predictions of peak width. The results show that the SDM is sufficient to describe the binding equilibrium of fusion proteins in ion-exchange columns packed with large resin particles. In addition, the binding mechanism between fusion proteins and the ion exchanger is explored with the assistance of comparative molecular modeling.
Zhang
Chenming
C
Department of Biological Systems Engineering, Virginia Polytechnic Institute and State University, Blacksburg, VA 24061, USA. cmzhang@vt.edu
Glatz
Charles E
CE
D002853:Chromatography, Liquid / Q000379:methods*; D005966:Glucuronidase / Q000378:metabolism; D008956:Models, Chemical; D011993:Recombinant Fusion Proteins / Q000737:chemistry
D011993:Recombinant Fusion Proteins; D005966:Glucuronidase