title: Conformational detection of p53's oligomeric state by FlAsH Fluorescence.
abstract: The p53 tumor suppressor protein is a critical checkpoint in prevention of tumor formation, and the function of p53 is dependent on proper formation of the active tetramer. In vitro studies have shown that p53 binds DNA most efficiently as a tetramer, though inactive p53 is predicted to be monomeric in vivo. We demonstrate that FlAsH binding can be used to distinguish between oligomeric states of p53, providing a potential tool to explore p53 oligomerization in vivo. The FlAsH tetra-cysteine binding motif has been incorporated along the dimer and tetramer interfaces in the p53 tetramerization domain to create reporters for the dimeric and tetrameric states of p53, though the geometry of the four cysteines is critical for efficient FlAsH binding. Furthermore, we demonstrate that FlAsH binding can be used to monitor tetramer formation in real-time. These results demonstrate the potential for using FlAsH fluorescence to monitor protein-protein interactions in vivo.
authors.affiliation: Department of Chemistry and Biochemistry, Northern Arizona University, Flagstaff, AZ 86011-5698, USA.
authors.forename: Tawnya M
authors.initials: TM
authors.lastname: Webber
authors.forename: Andrew C
authors.initials: AC
authors.lastname: Allen
authors.forename: Wai Kit
authors.initials: WK
authors.lastname: Ma
authors.forename: Rhett G
authors.initials: RG
authors.lastname: Molloy
authors.forename: Charisse N
authors.initials: CN
authors.lastname: Kettelkamp
authors.forename: Caitlin A
authors.initials: CA
authors.lastname: Dow
authors.forename: Matthew J
authors.initials: MJ
authors.lastname: Gage
mesh_terms: D020816:Amino Acid Motifs; D004247:DNA / Q000737:chemistry; D005453:Fluorescence; D006801:Humans; D011487:Protein Conformation; D055503:Protein Multimerization; D013050:Spectrometry, Fluorescence / Q000379:methods*; D016159:Tumor Suppressor Protein p53 / Q000737:chemistry* / Q000235:genetics
chemical_list: D016159:Tumor Suppressor Protein p53; D004247:DNA