Fmoc-based chemical synthesis and selective binding to supercoiled DNA of the p53 C-terminal segment and its phosphorylated and acetylated derivatives.
The C-terminal domain of p53 comprises a linker, the tetramerization domain and the regulatory domain, and contains at least seven sites of potential post-translational modification. An improved strategy was developed for the synthesis of large peptides that contain phosphorylated amino acids and p53(303-393), a 91-amino acid peptide, and three post-translationally modified derivatives were synthesized through the sequential condensation of three partially protected segments. Peptide thiolesters were prepared using the sulfonamide-based 'safety-catch' resin approach and employing Fmoc-based solid-phase peptide synthesis. At the N-terminus of the middle building block, a photolabile protecting group, 3,4-dimethoxy-6-nitrobenzyloxycarbonyl, was incorporated to differentiate the N-terminal amino group from the side-chain amino groups. Two sequential couplings were accomplished following this protection strategy. The synthetic products, p53(303-393) and its phosphorylated or acetylated derivatives, exhibited the ability to bind specifically to supercoiled DNA, which is one of the characteristics of this domain.
Teruya
Kenta
K
National Cancer Institute, NIH, 37 Convent Drive, Bethesda, MD 20892, USA.
Murphy
Angela C
AC
Burlin
Tom
T
Appella
Ettore
E
Mazur
Sharlyn J
SJ
D000107:Acetylation; D000596:Amino Acids; D001667:Binding, Competitive; D004278:DNA, Superhelical / Q000737:chemistry / Q000378:metabolism*; D005449:Fluorenes / Q000737:chemistry*; D006801:Humans; D008969:Molecular Sequence Data; D010455:Peptides / Q000138:chemical synthesis* / Q000737:chemistry / Q000378:metabolism*; D010766:Phosphorylation; D016159:Tumor Suppressor Protein p53 / Q000737:chemistry* / Q000378:metabolism*
C496789:9-fluorenylmethoxycarbonyl; D000596:Amino Acids; D004278:DNA, Superhelical; D005449:Fluorenes; D010455:Peptides; D016159:Tumor Suppressor Protein p53