A phosphorylation tag for uranyl mediated protein purification and photo assisted tag removal

Most protein purification procedures include an affinity tag fused to either the N or C-terminal end of the protein of interest as well as a procedure for tag removal. Tag removal is not straightforward and especially tag removal from the C-terminal end is a challenge due to the characteristics of e...

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Published in:PloS one 2014-03, Vol.9 (3), p.e91138-e91138
Main Authors: Zhang, Qiang, Jørgensen, Thomas J D, Nielsen, Peter E, Møllegaard, Niels Erik
Format: Article
Language:English
Subjects:
Amino acids
Animals
Biology
Casein Kinase II - metabolism
Cattle
Chemistry
Chromatography
Electrophoresis, Polyacrylamide Gel
Enzymes
Escherichia coli
Escherichia coli - metabolism
Green Fluorescent Proteins - metabolism
Light
Phosphorylation
Physics
Protein binding
Recombinant Fusion Proteins - isolation & purification
Recombinant Fusion Proteins - metabolism
Recombinant proteins
Sepharose
Spectrometry, Mass, Electrospray Ionization
Uranium - metabolism
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description Most protein purification procedures include an affinity tag fused to either the N or C-terminal end of the protein of interest as well as a procedure for tag removal. Tag removal is not straightforward and especially tag removal from the C-terminal end is a challenge due to the characteristics of enzymes available for this purpose. In the present study, we demonstrate the utility of the divalent uranyl ion in a new procedure for protein purification and tag removal. By employment of a GFP (green florescence protein) recombinant protein we show that uranyl binding to a phosphorylated C-terminal tag enables target protein purification from an E. coli extract by immobilized uranyl affinity chromatography. Subsequently, the tag can be efficiently removed by UV-irradiation assisted uranyl photocleavage. We therefore suggest that the divalent uranyl ion (UO22+) may provide a dual function in protein purification and subsequent C-terminal tag removal procedures.
doi_str_mv 10.1371/journal.pone.0091138
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subjects Amino acids
Animals
Biology
Casein Kinase II - metabolism
Cattle
Chemistry
Chromatography
Electrophoresis, Polyacrylamide Gel
Enzymes
Escherichia coli
Escherichia coli - metabolism
Green Fluorescent Proteins - metabolism
Light
Phosphorylation
Physics
Protein binding
Recombinant Fusion Proteins - isolation & purification
Recombinant Fusion Proteins - metabolism
Recombinant proteins
Sepharose
Spectrometry, Mass, Electrospray Ionization
Uranium - metabolism
title A phosphorylation tag for uranyl mediated protein purification and photo assisted tag removal
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