Peptide Binding by a Fragment of Calmodulin Composed of Ef-hands 2 and 3.
From: Faculty of Pharmaceutical Sciences, Division of Biomolecular and Pharmaceutical Chemistry, University of British Columbia, Vancouver, British Columbia, Canada V6T 1Z3.
Biochemistry
- Publish Date: Jul 2007
- ISSN: 0006-2960
- Volume: 46
- Issue: 29
- Pages: 8525-36
- Medium: Print
- Language: English
- Citation (JAMA): Lakowski Ted M, Lee Gregory M, Lelj-Garolla Barbara, et al. Peptide Binding by a Fragment of Calmodulin Composed of Ef-hands 2 and 3.. Biochemistry Jul 2007;46:8525-36
Abstract
Calmodulin (CaM) is composed of two EF-hand domains tethered by a flexible linker. Upon Ca2+-binding, a fragment of CaM encompassing EF-hands 2 and 3 (CaM2/3; residues 46-113) folds into a structure remarkably similar to the N- and C-domains of CaM. In this study, we demonstrate that Ca2+-ligated CaM2/3 can also bind to a peptide representing the CaM-recognition sequence of skeletal muscle myosin light chain kinase (M13) with an equimolar stoichiometry and a dissociation constant of 0.40 +/- 0.05 microM. On the basis of an analytical ultracentrifugation measurement, the resulting complex exists as an equilibrium mixture of 2:2 heterotetrameric and 1:1 heterodimeric species. Chemical shift perturbation mapping indicates that, similar to CaM, the peptide associates with a hydrophobic groove crossing both EF-hands in CaM2/3. However, upon binding the M13 peptide, many residues in CaM2/3 yielded two equal intensity NMR signals with the same 15N relaxation properties. Thus, the 2:2 CaM2/3-M13 tetramer, which predominates under the conditions used for these studies, is asymmetric with each component adopting spectroscopically distinguishable conformations within the complex. CaM2/3 also weakly stimulates the phosphatase activity of calcineurin and inhibits stimulation by native CaM. These studies highlight the remarkable plasticity of EF-hand association and expand the diverse repertoire of mechanisms possible for CaM-target protein interactions.
Mesh Headings (Keywords): Binding Sites, Calcineurin, Calmodulin, EF Hand Motifs, Humans, Hydrophobicity, Magnetic Resonance Spectroscopy, Models, Molecular, Myosin-Light-Chain Kinase, Peptide Fragments, Protein Structure, Secondary
Check for Full Text / PubMed Unique Identifier (PMID): 17595060
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