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

Research output: Contribution to journalArticle

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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 - Jan 1 2014

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Dipeptides
Calmodulin
Vasodilator Agents
Computer Simulation
Calcium
Histidine
Tryptophan
Peptides
histidylphenylalanine
Protein Kinases
In Vitro Techniques
Hydrogen
Calcium-Calmodulin-Dependent Protein Kinase Type 2
Basic Amino Acids
Molecular Dynamics Simulation
Encapsulation
Skeleton
Stoichiometry
Free energy
Molecular dynamics

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Biophysics
  • Molecular Biology

Cite this

Inhibition of calcium-calmodulin complex formation by vasorelaxant basic dipeptides demonstrated by in vitro and in silico analyses. / Kumrungsee, Thanutchaporn; Saiki, Tomomi; Akiyama, Sayaka; Nakashima, Kentaro; Tanaka, Mitsuru; Kobayashi, Yutaro; Matsui, Toshiro.

In: Biochimica et Biophysica Acta - General Subjects, Vol. 1840, No. 10, 01.01.2014, p. 3073-3078.

Research output: Contribution to journalArticle

Kumrungsee, Thanutchaporn ; Saiki, Tomomi ; Akiyama, Sayaka ; Nakashima, Kentaro ; Tanaka, Mitsuru ; Kobayashi, Yutaro ; Matsui, Toshiro. / Inhibition of calcium-calmodulin complex formation by vasorelaxant basic dipeptides demonstrated by in vitro and in silico analyses. In: Biochimica et Biophysica Acta - General Subjects. 2014 ; Vol. 1840, No. 10. pp. 3073-3078.
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AU - Kumrungsee, Thanutchaporn

AU - Saiki, Tomomi

AU - Akiyama, Sayaka

AU - Nakashima, Kentaro

AU - Tanaka, Mitsuru

AU - Kobayashi, Yutaro

AU - Matsui, Toshiro

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N2 - 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.

AB - 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.

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