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Domain Substructure of HPV E6 Oncoprotein: Biophysical Characterization of the E6 C-Terminal DNA-Binding Domain

Nominé, Yves, Charbonnier, Sebastian, Ristriani, Tutik, Stier, Gunter, Masson, Murielle, Cavusoglu, Nukhet, Van Dorsselaer, Alain, Weiss, Étienne, Kieffer, Bruno, Travé, Gilles·2003·Biochemistry
DOI 10.1021/bi026980c
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Abstract

E6 is a viral oncoprotein implicated in cervical cancers, produced by high-risk human papillomaviruses (HPVs). Structural data concerning this protein are scarce due to the difficulty of producing recombinant E6. Recently, we described the expression and purification of a stable, folded, and biologically active HPV16 E6 mutant called E6 6C/6S. Here, we analyzed the domain substructure of this mutated E6. Nonspecific proteolysis of full-length E6 6C/6S (158 residues) yielded N-terminal and C-terminal fragments encompassing residues 7-83 and 87-158, respectively. The C-terminal fragment of residues 87-158 was cloned, overexpressed, and purified at concentrations as high as 1 mM. The purified domain retains the selective four-way DNA junction recognition activity of the full-length E6 protein. Using UV absorption, UV fluorescence, circular dichroism, and nuclear magnetic resonance, we show that the peptide is primarily monomeric and folded with equal proportions of alpha-helix and beta-sheet secondary structure.

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Cited by

Metadata

Authors
Nominé, Y., Charbonnier, S., Ristriani, T., Stier, G., Masson, M., Cavusoglu, N., Van Dorsselaer, A., Weiss, É., Kieffer, B., & Travé, G.
Affiliations
Laboratoire d'Immunotechnologie, UMR CNRS 7100, École Supérieure de Biotechnologie de Strasbourg, 67400 Illkirch, France, EMBL, Meyerhofstrasse 1, D-69117 Heidelberg, Germany, Laboratoire de RMN, UMR CNRS 7104, École Supérieure de Biotechnologie de Strasbourg, 67400 Illkirch, France, and Laboratoire de Spectrométrie de Masse Bio-Organique, UMR CNRS 7509, Faculté de Chimie, 67087 Strasbourg, France
Venue
Biochemistry
Publisher
American Chemical Society (ACS)
Published
2003-04-08
Volume
42
Issue
17
Pages
4909-4917
ISSN
  • 0006-2960
  • 1520-4995
DOI
10.1021/bi026980c
Cited by
49
References
0
Language
en

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