Canonical structural-binding modes in the calmodulin–target protein complexes

Abstract

Intracellular calcium sensor protein calmodulin (CaM) belongs to the large EF-hand protein superfamily. CaM shows a unique and not fully understood ability to bind to multiple targets, allows them to participate in a variety of regulatory processes. The protein has two approximately symmetrical globular domains (the N- and C-lobes). Analysis of the CaM-binding sites of target proteins showed that they have two hydrophobic ‘anchor’ amino acids separated by 10 to 17 residues. Consequently, several CaM-binding motifs: {1–10}, {1–11}, {1–13}, {1–14}, {1–16}, {1–17}, differing by the distance between the two anchor residues along the amino acid sequence, have been identified. Despite extensive structural information on the role of target–protein amino acid residues in the formation of complexes with CaM, much less is known about the role of amino acids from CaM contributing to these interactions. In this work, a quantitative analysis of the contact surfaces of CaM and target proteins has been carried out for 35 representative three-dimensional structures. It has been shown that, in addition to the two hydrophobic terminal residues of the target fragment, the interaction also involves residues that are 4 residues earlier in the sequence (binding mode {1–5}). It has also been found that the N- and C-lobes of CaM bind the {1–5} motif located at the ends of the target in a structurally identical manner. Methionine residues at positions 51 (corresponding to 124 in the C-lobe), 71 (144), and 72 (145) of the CaM amino acid sequence are key hydrophobic residues for this interaction. They are located at the N- and C-boundaries of the even EF-hand motifs. The hydrophobic core of CaM (‘Ф-quatrefoil’) consists of 10 amino acids in the N-lobe (and in the C-lobe): Phe₁₆ (Phe₈₉), Phe₁₉ (Phe₉₂), Ile₂₇ (Ile₁₀₀), Thr₂₉ (Ala₁₀₂), Leu₃₂ (Leu₁₀₅), Ile₅₂ (Ile₁₂₅), Val₅₅ (Ala₁₂₈), Ile₆₃ (Val₁₃₆), Phe₆₅ (Tyr₁₃₈), and Phe₆₈ (Phe₁₄₁) and do not intersect with the target-binding methionine residues. CaM belongs to the ‘dynamic’ group of EF-hand proteins, in which calcium and protein ligand binding causes only global conformational changes but does not alter the conservative ‘black’ and ‘grey’ clusters described in our earlier works (PLoS One. 2014; 9(10):e109287). The membership of CaM in the ‘dynamic’ group is determined by the triggering and protective methionine layer: Met₅₁ (Met₁₂₄), Met₇₁ (Met₁₄₄) and Met₇₂ (Met₁₄₅).

HIGHLIGHTS

• Interchain interactions in the unique 35 CaM complex structures were analyzed.

• Methionine amino acids of the N- and C-lobes of CaM form triggering and protective layers.

• Interactions of the target terminal residues with these methionine layers are structurally identical.

• CaM belonging to the ‘dynamic’ group is determined by the triggering and protective methionine layer.

Communicated by Ramaswamy H. Sarma

Keywords:

• Calcium
• EF-hand
• calmodulin–target complex
• structural motif
• Ф-quatrefoil
• ‘black’ and ‘grey’ clusters

Acknowledgments

We thank the Biocenter Finland Bioinformatics Network (Dr. Jukka Lehtonen) and CSC IT Center for Science for computational support for the project. The Structural Bioinformatics Laboratory is part of the Solution for Health strategic area of Åbo Akademi University and within the InFLAMES Flagship program on inflammation and infection, Åbo Akademi University and the University of Turku, funded by the Academy of Finland.

Disclosure statement

The authors declare no conflict of interest.

Additional information

Funding

The project was supported by the Sigrid Jusélius Foundation and Joe, Pentti and Tor Borg Memorial Fund (A.I.D. and M.S.J).