Structural changes in the C‐terminus of Ca2+‐bound rat S100B(ββ) upon binding to a peptide derived from the C‐terminal regulatory domain of p53

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Abstract
S100B(ββ) is a dimeric Ca2+‐binding protein that interacts with p53, inhibits its phosphorylation by protein kinase C (PKC) and promotes disassembly of the p53 tetramer. Likewise, a 22 residue peptide derived from the C‐terminal regulatory domain of p53 has been shown to interact with S100B(ββ) in a Ca2+‐dependent manner and inhibits its phosphorylation by PKC. Hence, structural studies of Ca2+‐loaded S100B(ββ) bound to the p53 peptide were initiated to characterize this interaction. Analysis of nuclear Overhauser effect (NOE) correlations, amide proton exchange rates, 3JNH‐Hα coupling constants, and chemical shift index data show that, like apo‐ and Ca2+‐bound S100B(ββ), S100B remains a dimer in the p53 peptide complex, and each subunit has four helices (helix 1, Glu2–Arg20; helix 2, Lys29–Asn38; helix 3, Gln50–Asp61; helix 4, Phe70–Phe87), four loops (loop 1, Glu21–His25; loop 2, Glu39–Glu49; loop 3, Glu62–Gly66; loop 4, Phe88–Glu91), and two β‐strands (β‐strand 1, Lys26–Lys28; β‐strand 2, Glu67–Asp69), which forms a short antiparallel β‐sheet. However, in the presence of the p53 peptide helix 4 is longer by five residues than in apo‐ or Ca2+‐bound S100B(ββ). Furthermore, the amide proton exchange rates in helix 3 (K55, V56, E58, T59, L60, D61) are significantly slower than those of Ca2+‐bound S100B(ββ). Together, these observations plus inter‐molecular NOE correlations between the p53 peptide and S100B(ββ) support the notion that the p53 peptide binds in a region of S100B(ββ), which includes residues in helix 2, helix 3, loop 2, and the C‐terminal loop, and that binding of the p53 peptide interacts with and induces the extension of helix 4.