Purification of poly-histidine-tagged proteins.
His-tagging is the most widespread and versatile strategy used to purify recombinant proteins for biochemical and structural studies. Recombinant DNA methods are first used to engineer the addition of a short tract of poly-histidine tag (His-tag) to the N terminus or C terminus of a target protein. The His-tag is then exploited to enable purification of the "tagged" protein by immobilised metal affinity chromatography (IMAC). Here, we describe efficient procedures for the isolation of highly purified His-tagged target proteins from an Escherichia coli host using IMAC.
School of Biotechnology and National Centre for Sensor Research, Dublin City University, Dublin, Ireland.
Sinéad T
ST
Loughran
Dermot
D
Walls
D015153:Blotting, Western; D002021:Buffers; D002846:Chromatography, Affinity / Q000379:methods*; D003101:Collodion / Q000737:chemistry; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics; D005374:Filtration; D006639:Histidine / Q000235:genetics* / Q000302:isolation & purification; D006801:Humans; D055767:Immobilized Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification; D015778:Minor Histocompatibility Antigens; D011489:Protein Denaturation; D058849:Protein Refolding; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000737:chemistry / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification; D011993:Recombinant Fusion Proteins / Q000737:chemistry* / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification*; D012394:Rosaniline Dyes / Q000737:chemistry; D012995:Solubility; D013194:Staining and Labeling
C096816:BCL2-related protein A1; D002021:Buffers; D055767:Immobilized Proteins; D015778:Minor Histocompatibility Antigens; D019253:Proto-Oncogene Proteins c-bcl-2; D011993:Recombinant Fusion Proteins; D012394:Rosaniline Dyes; C033223:polyhistidine; D006639:Histidine; C048139:Coomassie blue; D003101:Collodion