Recruitment of the transcriptional machinery through GAL11P: structure andinteractions of the GAL4 dimerization domain

The GAL4 dimerization domain (GAL4-dd) is a powerful transcriptional activa tor when tethered to DNA in a cell bearing a mutant of the GAL11 protein, n amed GAL11P. GAL11P (like GAL11) is a component of the RNA-polymerase II ho loenzyme. Nuclear magnetic resonance (NMR) studies of GAL4-dd revealed an e longated dimer structure with C-2 symmetry containing three helices that me diate dimerization via coiled-coil contacts. The two loops between the thre e coiled coils form mobile bulges causing a variation of twist angles betwe en 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. Alanin e-scanning mutagenesis of GAL4-dd showed that the NMR-derived GAL11P-bindin g face is crucial for the novel transcriptional activating function of the GAL4-dd on GAL11P interaction. The binding of GAL4 to GAL11P, although an a rtificial interaction, represents a unique structural motif for an activati ng region capable of binding to a single target to effect gene expression.