Solution structure determination and mutational analysis of the papillomavirus E6 interacting peptide of E6AP.
E6AP is a cellular protein that binds cancer-related papillomaviral E6 proteins. The E6 binding domain, called E6ap, is located on an 18-amino acid segment of E6AP. The corresponding peptide was synthesized and its structure determined by nuclear magnetic resonance spectroscopy. The overall structure of the peptide is helical. A consensus E6-binding sequence among different E6 interacting proteins contains three conserved hydrophobic residues. In the structure of the E6AP peptide, the three conserved leucines (Leu 9, Leu 12, and Leu 13) form a hydrophobic patch on one face of the alpha-helix. Substitution of any of these leucines with alanine abolished binding to E6 protein, indicating that the entire hydrophobic patch is necessary. Mutation of a glutamate to proline, but not alanine, also disrupted the interaction between E6 and E6AP protein, suggesting that the E6-binding motif of the E6AP protein must be helical when bound to E6. Comparison of the E6ap structure and mutational results with those of another E6-binding protein (E6BP/ERC-55) indicates the existence of a general E6-binding motif.
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Department of Biochemistry, Tufts University School of Medicine, 136 Harrison Avenue, Boston, Massachusetts 02111, USA.
Hong
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Wei
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Androphy
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Chen
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Baleja
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D000595:Amino Acid Sequence; D052688:Bovine papillomavirus 1 / Q000737:chemistry / Q000235:genetics; D002942:Circular Dichroism; D018360:Crystallography, X-Ray; D004252:DNA Mutational Analysis; D006801:Humans; D008025:Ligases / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D019906:Nuclear Magnetic Resonance, Biomolecular; D009856:Oncogene Proteins, Viral / Q000378:metabolism*; D027383:Papillomaviridae / Q000737:chemistry / Q000235:genetics; D011485:Protein Binding / Q000235:genetics; D012097:Repressor Proteins; D012996:Solutions; D044767:Ubiquitin-Protein Ligases
C068651:E6 protein, Human papillomavirus type 16; D009856:Oncogene Proteins, Viral; D012097:Repressor Proteins; D012996:Solutions; C059269:protein E6, Bovine papillomavirus; D044767:Ubiquitin-Protein Ligases; D008025:Ligases