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標題: Adsorption behaviors of recombinant proteins on hydroxyapatite-based immobilized metal affinity chromatographic adsorbents
作者: Lin, P.C.
Lin, S.C.
Hsu, W.H.
關鍵字: IMAC;adsorption isotherm;protein purification;d-glucosamine 2-epimerase;substrate-specificity;varying particle;structural basis;purification;acid;resolution;peptides;binding;ions
Project: Journal of the Chinese Institute of Chemical Engineers
期刊/報告no:: Journal of the Chinese Institute of Chemical Engineers, Volume 39, Issue 5, Page(s) 389-398.
The equilibrium adsorption of three homo-oligoemric model recombinant proteins containing up to 8 poly(histidine) affinity tags on a hydroxyapatite-based immobilized metal affinity chromatography (IMAC) adsrobernt is reported in this study. The experimental data are well fitted with the three-parameter Langmuir-Freudlich isotherm model, indicating the presencce of positive cooperativity for the adsorption of these model proteins. The maximum capacity and the binding affinity of the IMAC adsorbent for the model proteins are in principle dependent on the size and the number of affinity tags of proteins, respectively. The exceptionally high association constant of the octameric racemase, probably due to simultaneous multipoint attachment, make it difficult to elute racemase from the adsorbent. The adsoption isotherms under denaturing conditions are well fitted with the Langmuir model. Result of Scatchard analysis further suggest the homogeneous adsorption of the model proteins subunits under denaturing conditions. The binding capacitites and affinities of the adsorbent under denaturing conditions for the three unfolded protein subunits become essentially identical because the molecular size and number of polu(His) tags of the unfolded polypeptide chains of the three protein subunits are the same. The significant reduction in association constants under denaturing conditions suggests that high concentration of urea could interfere with the binding of proteins on the hydroxyapatite-based adsorbent. (c) 2008 Taiwan Institute of Chemical Engineers, Published by Elsevier B.V. All rights reserved.
ISSN: 0368-1653
DOI: 10.1016/j.jcice.2008.03.007
Appears in Collections:化學工程學系所

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