Article ID | Journal | Published Year | Pages | File Type |
---|---|---|---|---|
9894304 | Protein Expression and Purification | 2005 | 9 Pages |
Abstract
Recombinant Staphylococcus aureus elongation factor G (EF-G) is difficult to refold by dilution due to the formation of large amounts of misfolded structures. However, refolding of EF-G by adsorption to a chromatographic column packed with immobilized polyethylene glycol 20,000 (PEG 20Â K) followed by pulse elution with 8Â M urea resulted in 88% mass recovery and 80% of correctly refolded structure. The PEG 20Â K was coupled to brominated allyl group derivatized Sepharose High Performance to construct a mild hydrophobic adsorbent. Various other hydrophobic interaction adsorbents were also attempted to refold EF-G. However, ligands with high hydrophobicity tended to misfold EF-G, resulting in irreversible adsorption. Various solvents, detergents, and low temperature as well as 8Â M urea were tried to release bound EF-G. Only pulse elution with 8Â M urea was efficient. Urea concentrations favorable for efficiently refolding EF-G were investigated. Low urea concentration produced more misfolded structures.
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Authors
Jing-Jing Li, Musturi Venkataramana, Ai-Qing Wang, Suparna Sanyal, Jan-Christer Janson, Zhi-Guo Su,