A mutation study of the DNA binding domain of human papillomavirus type11 E2 protein.
A site-specific mutation study was performed on the C-terminal domain, containing a cloned DNA binding region, of the human papillomavirus type11 (HPV11) E2 protein to determine the specific properties of residues directly involved in the DNA binding. The effect of a point mutations on the DNA binding was assessed by means of a gel mobility shift assay. The mutagenesis was concentrated on the residues in the third helix from the N-terminal, that is known as the "recognition helix," in the crystal structure of the bovine papillomavirus (BPV) E2 protein. Most point mutations caused a great decrease in the DNA binding activity. The leucine repeat in the DNA binding region was proved not to be a leucine prerequisite, as the leucines could be substituted by valine without significant loss of the DNA binding ability. Substitution of Leu for Glu caused a significant decrease in the DNA binding, indicating that the hydrophobicity of the residue at this position is important. The results suggest that the individual contribution of each amino acid residue in the DNA binding region is essential for the DNA binding.
Matsumoto
T
T
Department of Molecular Biology, National Institute of Agrobiological Resources, Ibaraki. mat@abr.affrc.go.jp
Nakashima
N
N
Takase
K
K
Hirochika
H
H
Mizuno
H
H
D000595:Amino Acid Sequence; D001483:Base Sequence; D001665:Binding Sites; D017124:Conserved Sequence; D004247:DNA / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D004586:Electrophoresis / Q000379:methods; D004926:Escherichia coli / Q000235:genetics / Q000821:virology; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009841:Oligonucleotides / Q000737:chemistry / Q000378:metabolism; D009856:Oncogene Proteins, Viral / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D027383:Papillomaviridae / Q000737:chemistry*; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D013379:Substrate Specificity; D014764:Viral Proteins / Q000378:metabolism
D004268:DNA-Binding Proteins; C051550:E2 protein, Bovine papillomavirus; D009841:Oligonucleotides; D009856:Oncogene Proteins, Viral; D011994:Recombinant Proteins; D014764:Viral Proteins; D004247:DNA