A C-terminal hydrophobic, solvent-protected core and a flexible N-terminus are potentially required for human papillomavirus 18 E7 protein functionality.
The oncogenic potential of the high-risk human papillomavirus (HPV) relies on the expression of genes specifying the E7 and E6 proteins. To investigate further the variation in oligomeric structure that has been reported for different E7 proteins, an HPV-18 E7 cloned from a Hispanic woman with cervical intraepithelial neoplasia was purified to homogeneity most probably as a stable monomeric protein in aqueous solution. We determined that one zinc ion is present per HPV-18 E7 monomer by amino acid and inductively coupled plasma-atomic emission spectroscopy analysis. Intrinsic fluorescence and circular dichroism spectroscopic results indicate that the zinc ion is important for the correct folding and thermal stability of HPV-18 E7. Hydroxyl radical mediated protein footprinting coupled to mass spectrometry and other biochemical and biophysical data indicate that near the C-terminus, the four cysteines of the two Cys-X(2)-Cys motifs that are coordinated to the zinc ion form a solvent inaccessible core. The N-terminal LXCXE pRb binding motif region is hydroxyl radical accessible and conformationally flexible. Both factors, the relative flexibility of the pRb binding motif at the N-terminus and the C-terminal metal-binding hydrophobic solvent-protected core, combine together and facilitate the biological functions of HPV-18 E7.
Liu
Shuqing
S
Department of Biochemistry and Molecular Biology, Dalian Medical University, Dalian 116044, China.
Tian
Yuxiang
Y
Greenaway
Frederick T
FT
Sun
Ming-Zhong
MZ
D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D005260:Female; D052161:Human papillomavirus 18; D006801:Humans; D057927:Hydrophobic and Hydrophilic Interactions; D017665:Hydroxyl Radical / Q000378:metabolism; D008969:Molecular Sequence Data; D009856:Oncogene Proteins, Viral / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D017510:Protein Folding; D055503:Protein Multimerization; D055550:Protein Stability; D020836:Protein Structure, Quaternary; D040901:Proteomics; D012997:Solvents / Q000737:chemistry*; D013329:Structure-Activity Relationship; D013696:Temperature; D014867:Water / Q000737:chemistry; D015032:Zinc / Q000378:metabolism
D004268:DNA-Binding Proteins; C052633:E7 protein, Human papillomavirus type 18; D009856:Oncogene Proteins, Viral; D012997:Solvents; D014867:Water; D017665:Hydroxyl Radical; D015032:Zinc