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

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Title: In-house preparation of hydrogels for batch affinity purification of glutathione S-transferase tagged recombinant proteins
Author(s): Buhrman, Jason S.; Rayahin, Jamie E.; Köllmer, Melanie; Gemeinhart, Richard A.
Subject(s): Glutathione PEGDA Glutathione S-transferase Batch purification
Abstract: Background: Many 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. Results: Glutathione 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. Conclusions: GSH 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.
Issue Date: 2012-09
Publisher: BioMed Central
Citation Info: Buhrman JS, Rayahin JE, Kollmer M, Gemeinhart RA. In-house preparation of hydrogels for batch affinity purification of glutathione S-transferase tagged recombinant proteins. BMC biotechnology. 2012;12:63. doi: 10.1186/1472-6750-12-63
Type: Article
Description: © 2012 Buhrman et al.; licensee 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.
URI: http://hdl.handle.net/10027/10839
ISSN: 1472-6750
Sponsor: This investigation was funded in part by the National Institutes of Health through grants R01 NS055095 (RAG) and in a facility constructed with support from Research Facilities Improvement Program Grant C06 RR15482 from the National Center for Research Resources, NIH.
Date Available in INDIGO: 2013-12-06
 

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