|
rdf:type |
|
|
lifeskim:mentions |
|
|
pubmed:issue |
3
|
|
pubmed:dateCreated |
2000-9-6
|
|
pubmed:abstractText |
We have developed a modified method of immobilized metal-ion affinity chromatography (IMAC) that can be used for the purification of histidine-tagged proteins from conditioned medium containing free copper ions. Classical methods of IMAC purification, using resins such as Ni-NTA, have proven inefficient for this type of purification and require multiple steps due to the interference of divalent copper ions with the binding of His-tagged protein to the charged resin. In contrast, this modified IMAC procedure, using chelating Sepharose instead of Ni-NTA, enables efficient purification from copper-containing medium in a single step. This method appears to rely upon a preferential interaction of protein-copper complexes with immobilized chelating resin. We have utilized this method to purify active, His-tagged murine interleukin 12 from the conditioned medium of Drosophila S2 cells coexpressing recombinant p40 and His-tagged p35 subunits and for the purification of the extracellular domain of the erythropoietin receptor. This method should be applicable to the purification of a wide variety of His-tagged fusion proteins expressed in Drosophila cells and in other systems where free metal ions are present.
|
|
pubmed:language |
eng
|
|
pubmed:journal |
|
|
pubmed:citationSubset |
IM
|
|
pubmed:chemical |
http://linkedlifedata.com/resource/pubmed/chemical/Chelating Agents,
http://linkedlifedata.com/resource/pubmed/chemical/Copper Sulfate,
http://linkedlifedata.com/resource/pubmed/chemical/Culture Media, Conditioned,
http://linkedlifedata.com/resource/pubmed/chemical/Histidine,
http://linkedlifedata.com/resource/pubmed/chemical/Interferon-gamma,
http://linkedlifedata.com/resource/pubmed/chemical/Interleukin-12,
http://linkedlifedata.com/resource/pubmed/chemical/Molecular Probes,
http://linkedlifedata.com/resource/pubmed/chemical/Receptors, Erythropoietin,
http://linkedlifedata.com/resource/pubmed/chemical/Recombinant Fusion Proteins
|
|
pubmed:status |
MEDLINE
|
|
pubmed:month |
Aug
|
|
pubmed:issn |
1046-5928
|
|
pubmed:author |
|
|
pubmed:copyrightInfo |
Copyright 2000 Academic Press.
|
|
pubmed:issnType |
Print
|
|
pubmed:volume |
19
|
|
pubmed:owner |
NLM
|
|
pubmed:authorsComplete |
Y
|
|
pubmed:pagination |
362-8
|
|
pubmed:dateRevised |
2008-11-21
|
|
pubmed:meshHeading |
pubmed-meshheading:10910726-Animals,
pubmed-meshheading:10910726-Blotting, Western,
pubmed-meshheading:10910726-Cell Line,
pubmed-meshheading:10910726-Cells, Cultured,
pubmed-meshheading:10910726-Chelating Agents,
pubmed-meshheading:10910726-Chromatography, Affinity,
pubmed-meshheading:10910726-Cloning, Molecular,
pubmed-meshheading:10910726-Copper Sulfate,
pubmed-meshheading:10910726-Culture Media, Conditioned,
pubmed-meshheading:10910726-Drosophila,
pubmed-meshheading:10910726-Electrophoresis, Polyacrylamide Gel,
pubmed-meshheading:10910726-Gene Expression Regulation,
pubmed-meshheading:10910726-Histidine,
pubmed-meshheading:10910726-Interferon-gamma,
pubmed-meshheading:10910726-Interleukin-12,
pubmed-meshheading:10910726-Mice,
pubmed-meshheading:10910726-Mice, Inbred BALB C,
pubmed-meshheading:10910726-Molecular Probes,
pubmed-meshheading:10910726-Plasmids,
pubmed-meshheading:10910726-Receptors, Erythropoietin,
pubmed-meshheading:10910726-Recombinant Fusion Proteins,
pubmed-meshheading:10910726-Spleen
|
|
pubmed:year |
2000
|
|
pubmed:articleTitle |
A modified metal-ion affinity chromatography procedure for the purification of histidine-tagged recombinant proteins expressed in Drosophila S2 cells.
|
|
pubmed:affiliation |
Department of Protein Biochemistry, SmithKline Beecham Pharmaceuticals, King of Prussia, Pennsylvania 19406, USA.
|
|
pubmed:publicationType |
Journal Article
|
