Structure
Volume 30, Issue 6, 2 June 2022, Pages 813-827.e5
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Article
DEER experiments reveal fundamental differences between calmodulin complexes with IQ and MARCKS peptides in solution

https://doi.org/10.1016/j.str.2022.03.005Get rights and content
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Highlights

  • The target peptides MARCKS and IQ bind CaM both in its apo- and holo-states

  • In solution, the binding of MARCKS to holo-CAM does not generate a closed conformation

  • In solution, holo-CaM with bound IQ has a closed conformation, albeit heterogeneous

  • Gd(III)/nitroxide DEER distance restraints give a solution holo-CaM/IQ structural model

Summary

Calmodulin (CaM) is a calcium-binding protein that regulates the function of many proteins by indirectly conferring Ca2+ sensitivity, and it undergoes a large conformational change on partners' binding. We compared the solution binding mode of the target peptides MARCKS and IQ by double electron-electron resonance (DEER) distance measurements and paramagnetic NMR. We combined nitroxide and Gd(III) spin labels, including specific substitution of one of the Ca2+ ions in the CaM mutant N60D by a Gd(III) ion. The binding of MARCKS to holo-CaM resulted neither in a closed conformation nor in a unique relative orientation between the two CaM domains, in contrast with the crystal structure. Binding of IQ to holo-CaM did generate a closed conformation. Using elastic network modeling and 12 distance restraints obtained from multiple holo-CaM/IQ DEER data, we derived a model of the solution structure, which is in reasonable agreement with the crystal structure.

Keywords

calmodulin
protein-peptide interaction
EPR
DEER
NMR
IQ-peptide
MARCKS
lanthanoid ions

Data and code availability

The primary DEER data and the restraints files used for the elastic network modeling were deposited in https://doi.org/10.5281/zenodo.6010570 and the NMR data were deposited in https://doi.org/10.5281/zenodo.5988755

This paper does not report original code.

Any additional information required to reanalyze the data reported in this paper is available from the lead contact upon request.

Cited by (0)

4

These authors contributed equally

5

Lead contact