The effects of CapZ peptide (TRTK-12) binding to S100B-Ca2+ as examined by NMR and X-ray crystallography.

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TitleThe effects of CapZ peptide (TRTK-12) binding to S100B-Ca2+ as examined by NMR and X-ray crystallography.
Publication TypeJournal Article
Year of Publication2010
AuthorsCharpentier, TH, Thompson, LE, Liriano, MA, Varney, KM, Wilder, PT, Pozharski, E, Toth, EA, Weber, DJ
JournalJ Mol Biol
Volume396
Issue5
Pagination1227-43
Date Published2010 Mar 12
ISSN1089-8638
KeywordsAmino Acid Sequence, Amino Acid Substitution, Animals, Binding Sites, Calcium, CapZ Actin Capping Protein, Cattle, Crystallography, X-Ray, Fluorescence Polarization, Humans, Kinetics, Models, Molecular, Multiprotein Complexes, Mutagenesis, Site-Directed, Nerve Growth Factors, Nuclear Magnetic Resonance, Biomolecular, Oligopeptides, Peptide Fragments, Protein Binding, Protein Conformation, Recombinant Proteins, S100 Calcium Binding Protein beta Subunit, S100 Proteins, Thermodynamics
Abstract

Structure-based drug design is underway to inhibit the S100B-p53 interaction as a strategy for treating malignant melanoma. X-ray crystallography was used here to characterize an interaction between Ca(2)(+)-S100B and TRTK-12, a target that binds to the p53-binding site on S100B. The structures of Ca(2+)-S100B (1.5-A resolution) and S100B-Ca(2)(+)-TRTK-12 (2.0-A resolution) determined here indicate that the S100B-Ca(2+)-TRTK-12 complex is dominated by an interaction between Trp7 of TRTK-12 and a hydrophobic binding pocket exposed on Ca(2+)-S100B involving residues in helices 2 and 3 and loop 2. As with an S100B-Ca(2)(+)-p53 peptide complex, TRTK-12 binding to Ca(2+)-S100B was found to increase the protein's Ca(2)(+)-binding affinity. One explanation for this effect was that peptide binding introduced a structural change that increased the number of Ca(2+) ligands and/or improved the Ca(2+) coordination geometry of S100B. This possibility was ruled out when the structures of S100B-Ca(2+)-TRTK-12 and S100B-Ca(2+) were compared and calcium ion coordination by the protein was found to be nearly identical in both EF-hand calcium-binding domains (RMSD=0.19). On the other hand, B-factors for residues in EF2 of Ca(2+)-S100B were found to be significantly lowered with TRTK-12 bound. This result is consistent with NMR (15)N relaxation studies that showed that TRTK-12 binding eliminated dynamic properties observed in Ca(2+)-S100B. Such a loss of protein motion may also provide an explanation for how calcium-ion-binding affinity is increased upon binding a target. Lastly, it follows that any small-molecule inhibitor bound to Ca(2+)-S100B would also have to cause an increase in calcium-ion-binding affinity to be effective therapeutically inside a cell, so these data need to be considered in future drug design studies involving S100B.

DOI10.1016/j.jmb.2009.12.057
Alternate JournalJ. Mol. Biol.
PubMed ID20053360
PubMed Central IDPMC2843395
Grant ListCA107331 / CA / NCI NIH HHS / United States
GM58888 / GM / NIGMS NIH HHS / United States
R01 CA107331-01A3 / CA / NCI NIH HHS / United States
R01 CA107331-02 / CA / NCI NIH HHS / United States
R01 CA107331-03 / CA / NCI NIH HHS / United States
R01 CA107331-04 / CA / NCI NIH HHS / United States
R01 CA107331-04S1 / CA / NCI NIH HHS / United States
R01 GM058888-08 / GM / NIGMS NIH HHS / United States
R01 GM058888-09 / GM / NIGMS NIH HHS / United States
R01 GM058888-10 / GM / NIGMS NIH HHS / United States
R01 GM058888-11 / GM / NIGMS NIH HHS / United States
R01 GM058888-11S1 / GM / NIGMS NIH HHS / United States
S10 RR015741-01 / RR / NCRR NIH HHS / United States
S10 RR016812-01 / RR / NCRR NIH HHS / United States
S10 RR023447-01 / RR / NCRR NIH HHS / United States