A physico-chemical investigation of the self-association of the DNA binding domain of the yeast transcriptional activator GAL4.
It has previously been suggested that the DNA binding domain (residues 1 to 147) of the yeast transcriptional activator GAL4 exists in solution in dimeric form, with the region responsible for dimerisation somewhere between residues 74 and 147. In this study limited proteolysis and carboxy-terminal deletions of the DNA binding domain (residues 1 to 147) of the yeast transcriptional activator GAL4 followed by subsequent characterization by equilibrium sedimentation in the analytical ultracentrifuge have been used to define more precisely the regions required for DNA binding and protein self-association. Sedimentation equilibrium analyses confirmed that the 'hydrophobic region' of the protein (residues 54-97, which contains a larger proportion of alpha-helix), is essential for dimerisation, with an apparent dissociation constant K(D,app), of approximately 50 microM for the 1-94 residue peptide and approximately 20 microM for the 1-147 residue peptide. Our studies do not rule out the possible formation of small amounts of additional higher order complexes.
Gadhavi
P
P
Department of Biochemistry, University of Cambridge, UK.
Morgan
P J
PJ
Alefounder
P
P
Harding
S E
SE
D000595:Amino Acid Sequence; D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D055592:Biophysical Phenomena; D001703:Biophysics; D055598:Chemical Phenomena; D002627:Chemistry, Physical; D004271:DNA, Fungal / Q000235:genetics / Q000378:metabolism*; D004268:DNA-Binding Proteins; D010450:Endopeptidases; D005656:Fungal Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins; D015534:Trans-Activators / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D014157:Transcription Factors
D004271:DNA, Fungal; D004268:DNA-Binding Proteins; D005656:Fungal Proteins; C052130:GAL4 protein, S cerevisiae; D010446:Peptide Fragments; D011994:Recombinant Proteins; D029701:Saccharomyces cerevisiae Proteins; D015534:Trans-Activators; D014157:Transcription Factors; D010450:Endopeptidases