In-house preparation of hydrogels for batch affinity purification of glutathione S-transferase tagged recombinant proteins

BMC Biotechnology

Methodology Article

Jamie E Rayahin¹  Melanie Köllmer¹  Jason S Buhrman¹  Richard A Gemeinhart² 

[1] Department of Biopharmaceutical Sciences, University of Illinois, 60612-7231, Chicago, IL, USA;Department of Biopharmaceutical Sciences, University of Illinois, 60612-7231, Chicago, IL, USA;Department of Bioengineering, University of Illinois, 60607-7052, Chicago, IL, USA;Department of Ophthalmology and Visual Science, University of Illinois, 60612-4319, Chicago, IL, USA;

关键词: Glutathione;    PEGDA;    Glutathione S-transferase;    Batch purification;    Recombinant protein;   

DOI  :  10.1186/1472-6750-12-63

 received in 2012-06-06, accepted in 2012-09-17,  发布年份 2012

来源: Springer

【 摘 要 】

BackgroundMany branches of biomedical research find use for pure recombinant proteins for direct application or to study other molecules and pathways. Glutathione affinity purification is commonly used to isolate and purify glutathione S-transferase (GST)-tagged fusion proteins from total cellular proteins in lysates. Although GST affinity materials are commercially available as glutathione immobilized on beaded agarose resins, few simple options for in-house production of those systems exist. Herein, we describe a novel method for the purification of GST-tagged recombinant proteins.ResultsGlutathione was conjugated to low molecular weight poly(ethylene glycol) diacrylate (PEGDA) via thiol-ene “click” chemistry. With our in-house prepared PEGDA:glutathione (PEGDA:GSH) homogenates, we were able to purify a glutathione S-transferase (GST) green fluorescent protein (GFP) fusion protein (GST-GFP) from the soluble fraction of E. coli lysate. Further, microspheres were formed from the PEGDA:GSH hydrogels and improved protein binding to a level comparable to purchased GSH-agarose beads.ConclusionsGSH containing polymers might find use as in-house methods of protein purification. They exhibited similar ability to purify GST tagged proteins as purchased GSH agarose beads.

【 授权许可】

Unknown   
© Buhrman et al.; licensee BioMed Central Ltd. 2012. This article is published under license to BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.