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Jungbauer, A; Machold, C; Hahn, R.
(2005): Hydrophobic interaction chromatography of proteins. III. Unfolding of proteins upon adsorption.
J Chromatogr A. 2005; 1079(1-2):221-228 FullText FullText_BOKU

Abstract:: Hydrophobic interaction chromatography (HIC) exploits the hydrophobic properties of protein surfaces for separation and purification by performing interactions with chromatographic sorbents of hydrophobic nature. In contrast to reversed-phase chromatography, this methodology is less detrimental to the protein and is therefore more commonly used in industrial scale as well as in bench scale when the conformational integrity of the protein is important. Hydrophobic interactions are promoted by salt and thus proteins are retained in presence of a cosmotropic salt. When proteins are injected on HIC columns with increasing salt concentrations under isocratic conditions only, a fraction of the applied amount is eluted. The higher the salt concentration, the lower is the amount of eluted protein. The rest can be desorbed with a buffer of low salt concentration or water. It has been proposed that the stronger retained protein fraction has partially changed the conformation upon adsorption. This has been also corroborated by physicochemical measurements. The retention data of 5 different model proteins and 10 different stationary phases were evaluated. Partial unfolding of proteins upon adsorption on surfaces of HIC media were assumed and a model describing the adsorption of native and partial unfolded fraction was developed. Furthermore, we hypothesize that the surface acts as catalyst for partial unfolding, since the fraction of partial unfolded protein is increasing with length of the alkyl chain. (c) 2005 Elsevier B.V. All rights reserved.
Authors BOKU Wien:: Hahn Rainer; Jungbauer Alois
Find related publications in this database (using NML MeSH Indexing): Adsorption -; Ammonium Sulfate - chemistry; Buffers -; Chromatography, Liquid -; Hydrophobicity -; Osmolar Concentration -; Protein Conformation -; Protein Denaturation -; Proteins - chemistry
Find related publications in this database (Keywords): protein; unfolding; pulse response; Langmuir-Freundlich isotherm; hydrophobic interaction