Effects of DNA binding and metal substitution on the dynamics of the GAL4 DNA-binding domain as studied by amide proton exchange.
Backbone amide proton exchange rates in the DNA-binding domain of GAL4 have been determined using 1H-15N heteronuclear correlation NMR spectroscopy. Three forms of the protein were studied-the native Zn-containing protein, the Cd-substituted protein, and a Zn-GAL4/DNA complex. Exchange rates in the Zn-containing protein are significantly slower than in the Cd-substituted protein. This shows that Cd-substituted GAL4 is destabilized relative to the native Zn-containing protein. Upon DNA binding, global retardation of amide proton exchange with solvent was observed, indicating that internal fluctuations of the DNA-recognition module are significantly reduced by the presence of DNA. In all forms of the protein, the internal dyad symmetry of the DNA-recognition module of GAL4 is reflected by the backbone amide proton exchange rates.
Mau
T
T
Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, Massachusetts 02115.
Baleja
J D
JD
Wagner
G
G
D000595:Amino Acid Sequence; D001483:Base Sequence; D004247:DNA / Q000737:chemistry / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics; D005656:Fungal Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D009682:Magnetic Resonance Spectroscopy / Q000379:methods; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides / Q000737:chemistry / Q000378:metabolism*; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors
D004268:DNA-Binding Proteins; D005656:Fungal Proteins; C052130:GAL4 protein, S cerevisiae; D009838:Oligodeoxyribonucleotides; D011994:Recombinant Proteins; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors; D004247:DNA