Expression and purification of human heat-shock transcription factor 1.
Heat-shock factor 1 (HSF1) is a 57-kDa cytoplasmic protein which binds to the promoters of heat-shock genes and activates transcription during heat shock. We describe here the expression and purification of the 529 amino acids form of human HSF1. We designed a new and complete purification protocol involving ammonium sulfate precipitation, heparin-Sepharose affinity, and ion-exchange chromatography, which allows the purification of large amounts of pure and active recombinant protein. HSF1 isolated by this method is pure as that assessed by SDS-PAGE and reverse-phase HPLC. The purified protein is recognized by specific anti-HSF1 antibodies, binds to heat-shock elements, and activates the promoter of the heat-shock protein 70A gene in vitro.
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Joint Center for Radiation Therapy, Harvard Medical School, Dana Farber Cancer Institute, Boston, Massachusetts 02115, USA.
Prevelige
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Calderwood
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D003001:Cloning, Molecular; D004268:DNA-Binding Proteins / Q000235:genetics / Q000302:isolation & purification*; D005786:Gene Expression Regulation; D018840:HSP70 Heat-Shock Proteins / Q000096:biosynthesis; D000076249:Heat Shock Transcription Factors; D006360:Heat-Shock Proteins / Q000235:genetics / Q000302:isolation & purification*; D006801:Humans; D011485:Protein Binding; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000302:isolation & purification; D012045:Regulatory Sequences, Nucleic Acid; D014157:Transcription Factors / Q000235:genetics / Q000302:isolation & purification*; D014158:Transcription, Genetic
D004268:DNA-Binding Proteins; C000619706:HSF1 protein, human; D018840:HSP70 Heat-Shock Proteins; D000076249:Heat Shock Transcription Factors; D006360:Heat-Shock Proteins; D011994:Recombinant Proteins; D014157:Transcription Factors