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

J Mol Biol. 2010 Mar 12;396(5):1227-43. doi: 10.1016/j.jmb.2009.12.057. Epub 2010 Jan 4.

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.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Amino Acid Substitution
  • Animals
  • Binding Sites
  • Calcium / metabolism*
  • CapZ Actin Capping Protein
  • Cattle
  • Crystallography, X-Ray
  • Fluorescence Polarization
  • Humans
  • In Vitro Techniques
  • Kinetics
  • Models, Molecular
  • Multiprotein Complexes
  • Mutagenesis, Site-Directed
  • Nerve Growth Factors / chemistry*
  • Nerve Growth Factors / genetics
  • Nerve Growth Factors / metabolism*
  • Nuclear Magnetic Resonance, Biomolecular
  • Oligopeptides / chemistry*
  • Oligopeptides / genetics
  • Oligopeptides / metabolism*
  • Peptide Fragments
  • Protein Binding
  • Protein Conformation
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / genetics
  • Recombinant Proteins / metabolism
  • S100 Calcium Binding Protein beta Subunit
  • S100 Proteins / chemistry*
  • S100 Proteins / genetics
  • S100 Proteins / metabolism*
  • Thermodynamics

Substances

  • CapZ Actin Capping Protein
  • Multiprotein Complexes
  • Nerve Growth Factors
  • Oligopeptides
  • Peptide Fragments
  • Recombinant Proteins
  • S100 Calcium Binding Protein beta Subunit
  • S100 Proteins
  • S100B protein, human
  • TRTK-12 peptide
  • Calcium

Associated data

  • PDB/3IQO
  • PDB/3IQQ