Human glutaredoxin 2 affinity tag for recombinant peptide and protein purification.
Recombinant proteins are commonly expressed in fusion with an affinity tag to facilitate purification. We have in the present study evaluated the possible use of the human glutaredoxin 2 (Grx2) as an affinity tag for purification of heterologous proteins. Grx2 is a glutathione binding protein and we have shown in the present study that the protein can be purified from crude bacterial extracts by a one-step affinity chromatography on glutathione-Sepharose. We further showed that short peptides could be fused to either the N- or C-terminus of Grx2 without affecting its ability to bind to the glutathione column. However, when Grx2 was fused to either the 27 kDa green fluorescent protein or the 116 kDa beta-galactosidase, the fusion proteins lost their ability to bind glutathione-Sepharose. Insertion of linker sequences between the Grx2 and the fusion protein did not restore binding to the column. In summary, our findings suggest that Grx2 may be used as an affinity tag for purification of short peptides and possibly also certain proteins that do not interfere with the binding to glutathione-Sepharose. However, the failure of purifying either green fluorescent protein or beta-galactosidase fused to Grx2 suggests that the use of Grx2 as an affinity tag for recombinant protein purification is limited.
Lundberg
Mathias
M
Karolinska Institute, Division of Clinical Virology, Karolinska University Hospital at Huddinge, S-141 86 Stockholm, Sweden.
Holmgren
Arne
A
Johansson
Magnus
M
D002846:Chromatography, Affinity; D003001:Cloning, Molecular; D005786:Gene Expression Regulation; D005822:Genetic Vectors / Q000096:biosynthesis / Q000235:genetics; D054477:Glutaredoxins; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000302:isolation & purification; D006801:Humans; D010088:Oxidoreductases / Q000235:genetics* / Q000302:isolation & purification*; D010446:Peptide Fragments / Q000235:genetics / Q000302:isolation & purification*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000302:isolation & purification; D001616:beta-Galactosidase / Q000235:genetics / Q000302:isolation & purification
C516008:GLRX2 protein, human; D054477:Glutaredoxins; D010446:Peptide Fragments; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins; D010088:Oxidoreductases; D001616:beta-Galactosidase