title: The GCN4 leucine zipper can functionally substitute for the heat shock transcription factor's trimerization domain.
abstract: The heat shock transcription factor (HSF) is the only known sequence-specific, homotrimeric DNA-binding protein. HSF binds to a DNA recognition site called a heat shock element (HSE), which contains varying numbers of nGAAn units ("GAA boxes") arranged in inverted repeats. To investigate the role of trimerization on HSF's DNA-binding properties, we replaced the trimerization domain, which self-assembles to form a three-stranded alpha-helical coiled coil, with the GCN4 leucine zipper, which forms a two-stranded alpha-helical coiled coil. Surprisingly, this substitution did not effect the ability of HSF to function in vivo. Biochemical studies of an HSF-leucine zipper chimera in comparison to an HSF truncation show that the HSF-leucine zipper chimera, though dimeric in solution and dimeric when bound to a two-box HSE, forms a trimeric complex when bound to a three-box HSE. The ability to form trimers depends on the presence of three contiguous GAA boxes present in inverted repeats. The proximity of the leucine zippers due to the orientation of the binding sites suggests that the leucine zippers might be forming a three-stranded coiled coil and several experiments lend support to this model. The ability of the leucine zipper to change oligomeric states in context might explain why the leucine zipper can replace the trimerization domain of HSF in vivo.
authors.affiliation: University of California, Department of Molecular and Cell Biology, Berkeley 94720-3206, USA.
authors.forename: B L
authors.initials: BL
authors.lastname: Drees
authors.forename: E K
authors.initials: EK
authors.lastname: Grotkopp
authors.forename: H C
authors.initials: HC
authors.lastname: Nelson
mesh_terms: D001665:Binding Sites; D003432:Cross-Linking Reagents / Q000378:metabolism; D004247:DNA / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics; D005656:Fungal Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D015870:Gene Expression; D000076249:Heat Shock Transcription Factors; D016350:Leucine Zippers; D008958:Models, Molecular; D009690:Nucleic Acid Conformation; D017354:Point Mutation; D011487:Protein Conformation; D011494:Protein Kinases / Q000737:chemistry / Q000378:metabolism*; D017433:Protein Structure, Secondary; D011993:Recombinant Fusion Proteins / Q000737:chemistry; D012441:Saccharomyces cerevisiae / Q000737:chemistry; D029701:Saccharomyces cerevisiae Proteins; D013388:Succinimides / Q000378:metabolism; D014157:Transcription Factors / Q000737:chemistry* / Q000378:metabolism
chemical_list: D003432:Cross-Linking Reagents; D004268:DNA-Binding Proteins; D005656:Fungal Proteins; D000076249:Heat Shock Transcription Factors; D011993:Recombinant Fusion Proteins; D029701:Saccharomyces cerevisiae Proteins; D013388:Succinimides; D014157:Transcription Factors; C028722:ethylene glycolylbis(succinimidyl succinate); D004247:DNA; D011494:Protein Kinases