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


Autoria(s): Forest, Amelie; Swulius, Matthew T; Tse, Joyce K Y; Bradshaw, J Michael; Gaertner, Tara; Waxham, M Neal
Data(s)

07/10/2008

Resumo

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 Ca (2+)-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 Ca (2+)-binding to each lobe of CaM during intracellular Ca (2+) oscillations.

Identificador

http://digitalcommons.library.tmc.edu/uthmed_docs/317

http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2665295/?tool=pmcentrez

Publicador

DigitalCommons@The Texas Medical Center

Fonte

UT Medical School Journal Articles

Palavras-Chave #Adenosine Diphosphate #Animals #Binding Sites #Calcium #Calcium-Calmodulin-Dependent Protein Kinase Type 2 #Calmodulin #Calorimetry #Fluorometry #Models #Molecular #Nucleotides #Phosphorylation #Protein Binding #Protein Structure #Tertiary #Rats #Temperature #Models, Molecular #Protein Structure, Tertiary #Medicine and Health Sciences
Tipo

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