Expression and Purification of Calmodulin for NMR and Other Biophysical Applications

  • Benjamin M. M. Grant
  • Christopher B. MarshallEmail author
  • Mitsuhiko IkuraEmail author
Part of the Methods in Molecular Biology book series (MIMB, volume 1929)


Calmodulin (CaM) is a ubiquitous calcium-sensing protein that has one of the most highly conserved sequences among eukaryotes. CaM has been a useful tool for biologists studying calcium signaling for decades. In recent years, CaM has also been implicated in numerous cancer-associated pathways, and rare CaM mutations have been identified as a cause of human cardiac arrhythmias. Here, we present a collection of our most recent and effective protocols for the expression and purification of recombinant CaM from Escherichia coli, including various isotopic labeling schemes, primarily for nuclear magnetic resonance (NMR) spectroscopy and other biophysical applications.

Key words

Calmodulin CaM Calcium Expression Purification NMR Isotopic labeling 



This work was supported by grants from Canadian Institutes of Health Research, Natural Sciences and Engineering Research Council of Canada, and Heart and Stroke Foundation of Canada. M.I. holds the Canada Research Chair in Cancer Structural Biology. The NMR facility at UHN is supported by Canada Foundation for Innovation and the Princess Margaret Foundation.

This article is in memory of Dr. Claude B. Klee for her encouragement and support of MI.


  1. 1.
    Cheung WY (1970) Cyclic 3′ -5′-nucleotide phosphodiesterase. Demonstration of an activator. Biochem Biophys Res Commun 38(3):533–538CrossRefGoogle Scholar
  2. 2.
    Kakiuchi S, Yamazaki R (1970) Calcium dependent phosphodiesterase activity and its activating factor (PAF) from brain studies on cyclic 3′,5′-nucleotide phosphodiesterase. Biochem Biophys Res Commun 41(5):1104–1110CrossRefGoogle Scholar
  3. 3.
    Cohen P, Klee CB (eds) (1988) Calmodulin. Molecular aspects of cellular regulation, vol Vol. 5. Elsevier, AmsterdamGoogle Scholar
  4. 4.
    Marshall CB et al (2015) Calmodulin and STIM proteins: two major calcium sensors in the cytoplasm and endoplasmic reticulum. Biochem Biophys Res Commun 460(1):5–21CrossRefGoogle Scholar
  5. 5.
    O’Connell DBM, Marshall CB, Ikura M, Linse S (2013) Calmodulin. In: Kretsinger RH, Uversky VN, Permyakov EA (eds) Encyclopedia of metalloproteins. Springer, BerlinGoogle Scholar
  6. 6.
    Moews PC, Kretsinger RH (1975) Terbium replacement of calcium in carp muscle calcium-binding parvalbumin: an x-ray crystallographic study. J Mol Biol 91(2):229–232CrossRefGoogle Scholar
  7. 7.
    Kawasaki H, Kretsinger RH (2017) Structural and functional diversity of EF-hand proteins: Evolutionary perspectives. Protein Sci 26:1898–1920CrossRefGoogle Scholar
  8. 8.
    Ikura M, Kay LE, Bax A (1990) Novel approach for sequential assignment of 1H, 13C, 15N spectra of proteins: heteronuclear triple-resonance three-dimensional NMR spectroscopy. Application to calmodulin. Biochemistry 29(19):4659–4667CrossRefGoogle Scholar
  9. 9.
    Ikura M, Kay LE, Bax A (1991) Improved three-dimensional 1H-13C-1H correlation spectroscopy of a 13C-labeled protein using constant-time evolution. J Biomol NMR 1(3):299–304CrossRefGoogle Scholar
  10. 10.
    Ikura M, Spera S, Barbato G, Kay LE, Krinks M, Bax A (1991) Secondary structure and side-chain 1H and 13C resonance assignments of calmodulin in solution by heteronuclear multidimensional NMR spectroscopy. Biochemistry 30(38):9216–9228CrossRefGoogle Scholar
  11. 11.
    Muallem S, Karlish SJ (1979) A simple, rapid and efficient procedure for purification of calmodulin from human red blood cells. FEBS Lett 107(1):209–212CrossRefGoogle Scholar
  12. 12.
    Yoshida M, Minowa O, Yagi K (1983) Divalent cation binding to wheat germ calmodulin. J Biochem 94(6):1925–1933CrossRefGoogle Scholar
  13. 13.
    Ikura M, Marion D, Kay LE, Shih H, Krinks M, Klee CB, Bax A (1990) Heteronuclear 3D NMR and isotopic labeling of calmodulin. Towards the complete assignment of the 1H NMR spectrum. Biochem Pharmacol 40(1):153–160CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  1. 1.Department of Medical BiophysicsUniversity of TorontoTorontoCanada
  2. 2.Princess Margaret Cancer CentreUniversity Health NetworkTorontoCanada

Personalised recommendations