The single Cys2-His2 zinc finger domain of the GAGA protein flanked by basic residues is sufficient for high-affinity specific DNA binding.
Specific DNA binding to the core consensus site GAGAGAG has been shown with an 82-residue peptide (residues 310-391) taken from the Drosophila transcription factor GAGA. Using a series of deletion mutants, it was demonstrated that the minimal domain required for specific binding (residues 310-372) includes a single zinc finger of the Cys2-His2 family and a stretch of basic amino acids located on the N-terminal end of the zinc finger. In gel retardation assays, the specific binding seen with either the peptide or the whole protein is zinc dependent and corresponds to a dissociation constant of approximately 5 x 10(-9) M for the purified peptide. It has previously been thought that a single zinc finger of the Cys2-His2 family is incapable of specific, high-affinity binding to DNA. The combination of an N-terminal basic region with a single Cys2-His2 zinc finger in the GAGA protein can thus be viewed as a novel DNA binding domain. This raises the possibility that other proteins carrying only one Cys2-His2 finger are also capable of high-affinity specific binding to DNA.
Pedone
P V
PV
Laboratory of Molecular Biology, Naional Institutes of Health, Bethesda, MD 20892-0530, USA.
Ghirlando
R
R
Clore
G M
GM
Gronenborn
A M
AM
Felsenfeld
G
G
Omichinski
J G
JG
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D002460:Cell Line; D003001:Cloning, Molecular; D004247:DNA / Q000378:metabolism*; D017931:DNA Primers; D004268:DNA-Binding Proteins; D004330:Drosophila / Q000378:metabolism; D029721:Drosophila Proteins; D018398:Homeodomain Proteins / Q000096:biosynthesis / Q000737:chemistry* / Q000378:metabolism*; D007700:Kinetics; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D016133:Polymerase Chain Reaction / Q000379:methods; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000378:metabolism; D014157:Transcription Factors / Q000096:biosynthesis / Q000737:chemistry* / Q000378:metabolism*; D014162:Transfection; D016335:Zinc Fingers
D017931:DNA Primers; D004268:DNA-Binding Proteins; D029721:Drosophila Proteins; D018398:Homeodomain Proteins; D010446:Peptide Fragments; D011994:Recombinant Proteins; D014157:Transcription Factors; C063460:Trl protein, Drosophila; D004247:DNA