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Related Concept Videos

¹H NMR of Conformationally Flexible Molecules: Variable-Temperature NMR01:15

¹H NMR of Conformationally Flexible Molecules: Variable-Temperature NMR

The axial and equatorial protons in cyclohexane can be distinguished by performing a variable-temperature NMR experiment. In this process, except for one proton, the remaining eleven protons are replaced by deuterium. The deuterium substitution avoids the possible peak splitting caused by the spin-spin coupling between the adjacent protons. The remaining proton flips between the axial and equatorial positions.
Calmodulin-dependent Signaling01:16

Calmodulin-dependent Signaling

Calmodulin (CaM) is a calcium-binding protein in eukaryotes that controls various calcium-regulated cellular processes. It has four calcium-binding sites that bind calcium to form the calcium-calmodulin ( Ca2+-CaM) complex. GPCR stimulation increases the calcium levels in the cells that bind to CaM and induces a conformational change.
The Ca2+-CaM complex does not have enzymatic activity by itself. Instead, the complex binds downstream target proteins, including membrane proteins or enzymes,...

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Related Experiment Video

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Pull-down of Calmodulin-binding Proteins
07:51

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Published on: January 23, 2012

Solution structure of a calmodulin-target peptide complex by multidimensional NMR.

M Ikura1, G M Clore, A M Gronenborn

  • 1Laboratory of Chemical Physics, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD 20892.

Science (New York, N.Y.)
|May 1, 1992
PubMed
Summary

Calcium-bound calmodulin (CaM) and a myosin light chain kinase peptide form a globular complex. Nuclear magnetic resonance reveals a disrupted central helix in CaM, allowing it to clamp the helical peptide via hydrophobic interactions.

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Area of Science:

  • Biochemistry
  • Structural Biology
  • Molecular Biology

Background:

  • Calmodulin (CaM) is a crucial calcium-binding protein involved in numerous cellular signaling pathways.
  • Skeletal muscle myosin light chain kinase (MLCK) contains a CaM-binding domain essential for regulating muscle contraction.
  • Understanding the structural basis of CaM-peptide interactions is key to deciphering cellular signaling mechanisms.

Purpose of the Study:

  • To determine the three-dimensional solution structure of the complex between calcium-bound calmodulin and a synthetic peptide representing the CaM-binding domain of skeletal muscle MLCK.
  • To elucidate the structural rearrangements in CaM upon peptide binding and the nature of the interactions stabilizing the complex.

Main Methods:

  • Multidimensional heteronuclear filtered and separated nuclear magnetic resonance (NMR) spectroscopy was employed to determine the solution structure.
  • Analysis of NMR data allowed for the characterization of CaM and peptide conformations and their spatial arrangement.

Main Results:

  • The Ca(2+)-CaM and MLCK peptide (residues 577-602) form a globular complex with overall dimensions of 47x32x30 angstroms.
  • CaM's central helix (residues 65-93) is disrupted into two helices connected by a flexible loop, enabling CaM's two domains to clamp the helical peptide (residues 3-21).
  • The helical peptide resides in a hydrophobic channel within the CaM ellipsoid, stabilized by extensive hydrophobic interactions, notably involving numerous methionine residues from CaM.

Conclusions:

  • The CaM-binding domain of MLCK adopts a helical conformation and binds to CaM through hydrophobic interactions, with key anchoring roles for Trp4 and Phe17 of the peptide.
  • CaM undergoes significant structural rearrangement, specifically the disruption of its central helix, to accommodate the binding peptide.
  • This study provides insights into a common binding mode for CaM-interacting peptides, suggesting a conserved mechanism involving aromatic and hydrophobic residues.