The homeodomain proteins, a1 and alpha 2, act cooperatively to regulate cell-type specific genes in yeast. The basis of this cooperativity is an interaction between the two proteins, forming a heterodimer that binds DNA tightly and specifically. A fragment containing the homeodomain of a1, a1(66-126), has been studied by NMR spectroscopy to gain secondary structure information and to characterize the changes in a1 upon heterodimerization with alpha 2. Heteronuclear (1H-15N) NMR methods were used to assign backbone resonances of the 61 amino acid fragment. The a1(66-126) secondary structure was determined using NOE connectivities, 3JHN alpha coupling constants and hydrogen exchange kinetic data. NMR data identify three helical segments separated by a loop and a tight turn that are the characteristic structural elements of homeodomain proteins. The a1 fragment was titrated with alpha 2(128-210), the homeodomain-containing fragment of alpha 2, to study changes in a1(66-126) spectra produced by alpha 2 binding. The a1(66-126) protein was labeled with 15N and selectively observed using isotope-edited NMR experiments. NMR spectra of bound a1(66-126) indicate that residues in helix 1, helix 2, and the loop connecting them are directly involved in the binding of the alpha 2 fragment. Relatively minor effects on the resonances from residues in helix 3, the putative DNA-binding helix, were noted upon alpha 2 binding. We have thus located a region of the a1 homeodomain important for specific protein recognition.
|Evidence ID||Analyze ID||Interactor||Interactor Systematic Name||Interactor||Interactor Systematic Name||Type||Assay||Annotation||Action||Modification||Phenotype||Source||Reference||Note|
|Evidence ID||Analyze ID||Gene||Gene Systematic Name||Gene Ontology Term||Gene Ontology Term ID||Qualifier||Aspect||Method||Evidence||Source||Assigned On||Reference||Annotation Extension|
|Evidence ID||Analyze ID||Gene||Gene Systematic Name||Phenotype||Experiment Type||Experiment Type Category||Mutant Information||Strain Background||Chemical||Details||Reference|
|Evidence ID||Analyze ID||Regulator||Regulator Systematic Name||Target||Target Systematic Name||Experiment||Conditions||Strain||Source||Reference|