title: Purification and characterization of a heat-shock element binding protein from yeast.
abstract: The promoters of heat shock genes are activated when cells are stressed. Activation is dependent on a specific DNA sequence, the heat-shock element (HSE). We describe the purification to homogeneity of an HSE-binding protein from yeast (Saccharomyces cerevisiae), using sequential chromatography of whole cell extracts on heparin-agarose, calf thymus DNA-Sepharose and an affinity column consisting of a repetitive synthetic HSE sequence coupled to Sepharose. The protein runs as a closely spaced doublet of approximately 150 kd on SDS-polyacrylamide gels; mild proteolysis generates a stable 70-kd fragment which retains DNA binding activity. The relative affinities of the protein for a range of variant HSE sequences correlates with the ability of these sequences to support heat-inducible transcription in vivo, suggesting that this polypeptide is involved in the activation of heat-shock promoters. However, the protein was purified from unshocked yeast, and may therefore represent an unactivated form of heat-shock transcription factor. Study of the purified protein should help to define the mechanistic basis of the heat-shock response.
authors.affiliation: MRC Laboratory of Molecular Biology, Cambridge, UK.
authors.forename: P K
authors.initials: PK
authors.lastname: Sorger
authors.forename: H R
authors.initials: HR
authors.lastname: Pelham
mesh_terms: D001483:Base Sequence; D004268:DNA-Binding Proteins / Q000302:isolation & purification / Q000378:metabolism*; D005796:Genes; D005800:Genes, Fungal; D014644:Genetic Variation; D006360:Heat-Shock Proteins / Q000235:genetics*; D010957:Plasmids; D011401:Promoter Regions, Genetic; D012441:Saccharomyces cerevisiae / Q000235:genetics*; D014158:Transcription, Genetic
chemical_list: D004268:DNA-Binding Proteins; D006360:Heat-Shock Proteins