Inhibition of calcium-calmodulin complex formation by vasorelaxant basic dipeptides demonstrated by in vitro and in silico analyses

Thanutchaporn Kumrungsee, Tomomi Saiki, Sayaka Akiyama, Kentaro Nakashima, Mitsuru Tanaka, Yutaro Kobayashi, Toshiro Matsui

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Abstract

Background Tryptophan-histidine (Trp-His) was found to suppress the activity of the Ca2 +/calmodulin (CaM)-dependent protein kinases II (CaMKII), which requires the Ca2 +-CaM complex for an initial activation. In this study, we attempted to clarify whether Trp-His inhibits Ca2 +-CaM complex formation, a CaMKII activator. Methods The ability of Trp-His and other peptides to inhibit Ca2 +-CaM complex formation was investigated by a Ca2 +-encapsulation fluorescence assay. The peptide-CaM interactions were illustrated by molecular dynamic simulation. Results We showed that Trp-His inhibited Ca2 +-CaM complex formation with a 1:1 binding stoichiometry of the peptide to CaM, considering that Trp-His reduced Hill coefficient of Ca2 +-CaM binding from 2.81 to 1.92. His-Trp also showed inhibitory activity, whereas Trp + His, 3-methyl His-Trp, and Phe-His did not show significant inhibitory activity, suggesting that the inhibitory activity was due to a peptide skeleton (irrespective of the sequence), a basic amino acid, a His residue, the N hydrogen atom of its imidazole ring, and Trp residue. In silico studies suggested the possibility that Trp-His and His-Trp interacted with the Ca2 +-binding site of CaM by forming hydrogen bonds with key Ca2 +-binding residues of CaM, with a binding free energy of - 49.1 and - 68.0 kJ/mol, respectively. Conclusions This is the first study demonstrating that the vasoactive dipeptide Trp-His possesses inhibitory activity against Ca2 +-CaM complex formation, which may elucidate how Trp-His inhibited CaMKII in a previous study. General significance The results provide a basic idea that could lead to the development of small peptides binding with high affinity to CaM and inhibiting Ca2 +-CaM complex formation in the future.

Original languageEnglish
Pages (from-to)3073-3078
Number of pages6
JournalBiochimica et Biophysica Acta - General Subjects
Volume1840
Issue number10
DOIs
Publication statusPublished - Oct 2014

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All Science Journal Classification (ASJC) codes

  • Biophysics
  • Biochemistry
  • Molecular Biology

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