title: Recruitment of the transcriptional machinery through GAL11P: structure and interactions of the GAL4 dimerization domain.
abstract: The GAL4 dimerization domain (GAL4-dd) is a powerful transcriptional activator when tethered to DNA in a cell bearing a mutant of the GAL11 protein, named GAL11P. GAL11P (like GAL11) is a component of the RNA-polymerase II holoenzyme. Nuclear magnetic resonance (NMR) studies of GAL4-dd revealed an elongated dimer structure with C(2) symmetry containing three helices that mediate dimerization via coiled-coil contacts. The two loops between the three coiled coils form mobile bulges causing a variation of twist angles between the helix pairs. Chemical shift perturbation analysis mapped the GAL11P-binding site to the C-terminal helix alpha3 and the loop between alpha1 and alpha2. One GAL11P monomer binds to one GAL4-dd dimer rendering the dimer asymmetric and implying an extreme negative cooperativity mechanism. Alanine-scanning mutagenesis of GAL4-dd showed that the NMR-derived GAL11P-binding face is crucial for the novel transcriptional activating function of the GAL4-dd on GAL11P interaction. The binding of GAL4 to GAL11P, although an artificial interaction, represents a unique structural motif for an activating region capable of binding to a single target to effect gene expression.
authors.affiliation: Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, Massachusetts 02115, USA.
authors.forename: P
authors.initials: P
authors.lastname: Hidalgo
authors.forename: A Z
authors.initials: AZ
authors.lastname: Ansari
authors.forename: P
authors.initials: P
authors.lastname: Schmidt
authors.forename: B
authors.initials: B
authors.lastname: Hare
authors.forename: N
authors.initials: N
authors.lastname: Simkovich
authors.forename: S
authors.initials: S
authors.lastname: Farrell
authors.forename: E J
authors.initials: EJ
authors.lastname: Shin
authors.forename: M
authors.initials: M
authors.lastname: Ptashne
authors.forename: G
authors.initials: G
authors.lastname: Wagner
mesh_terms: D000409:Alanine / Q000378:metabolism; D000595:Amino Acid Sequence; D001665:Binding Sites; D004247:DNA / Q000378:metabolism; D004268:DNA-Binding Proteins; D019281:Dimerization; D005656:Fungal Proteins / Q000737:chemistry* / Q000378:metabolism; D009682:Magnetic Resonance Spectroscopy; D056892:Mediator Complex; D008957:Models, Genetic; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D017510:Protein Folding; D017434:Protein Structure, Tertiary; D012319:RNA Polymerase II / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000737:chemistry; D029701:Saccharomyces cerevisiae Proteins; D017386:Sequence Homology, Amino Acid; D013696:Temperature; D015534:Trans-Activators / Q000737:chemistry* / Q000378:metabolism; D014157:Transcription Factors / Q000737:chemistry* / Q000378:metabolism; D014158:Transcription, Genetic
chemical_list: D004268:DNA-Binding Proteins; D005656:Fungal Proteins; C058337:GAL11 protein, S cerevisiae; C052130:GAL4 protein, S cerevisiae; D056892:Mediator Complex; D029701:Saccharomyces cerevisiae Proteins; D015534:Trans-Activators; D014157:Transcription Factors; D004247:DNA; D012319:RNA Polymerase II; D000409:Alanine