Role of the N- and C-lobes of calmodulin in the activation of Ca 2+/calmodulin-dependent protein kinase II

Amelie Forest, Matthew T. Swulius, Joyce K.Y. Tse, J. Michael Bradshaw, Tara Gaertner, M. Neal Waxham

Research output: Contribution to journalArticlepeer-review

28 Scopus citations

Abstract

Understanding the principles of calmodulin (CaM) activation of target enzymes will help delineate how this seemingly simple molecule can play such a complex role in transducing Ca2+-signals to a variety of downstream pathways. In the work reported here, we use biochemical and biophysical tools and a panel of CaM constructs to examine the lobe specific interactions between CaM and CaMKII necessary for the activation and autophosphorylation of the enzyme. Interestingly, the N-terminal lobe of CaM by itself was able to partially activate and allow autophosphorylation of CaMKII while the C-terminal lobe was inactive. When used together, CaMN and CaMC produced maximal CaMKII activation and autophosphorylation. Moreover, CaMNN and CaMCC (chimeras of the two N- or C-terminal lobes) both activated the kinase but with greater K act than for WtCaM. Isothermal titration calorimetry experiments showed the same rank order of affinities of WtCaM > CaMNN > CaMCC as those determined in the activity assay and that the CaM to CaMKII subunit binding ratio was 1:1. Together, our results lead to a proposed sequential mechanism to describe the activation pathway of CaMKII led by binding of the N-lobe followed by the C-lobe. This mechanism contrasts the typical sequential binding mode of CaM with other CaM-dependent enzymes, where the C-lobe of CaM binds first. The consequence of such lobe specific binding mechanisms is discussed in relation to the differential rates of Ca2+-binding to each lobe of CaM during intracellular Ca2+ oscillations.

Original languageEnglish (US)
Pages (from-to)10587-10599
Number of pages13
JournalBiochemistry
Volume47
Issue number40
DOIs
StatePublished - Oct 7 2008

All Science Journal Classification (ASJC) codes

  • Biochemistry

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