Expression of a multifunctional Ca2+/calmodulin-dependent protein kinase and mutational analysis of its autoregulation

doi:10.1016/0896-6273(89)90115-3

Neuron

Volume 3, Issue 1, July 1989, Pages 59-70

https://doi.org/10.1016/0896-6273(89)90115-3 Get rights and content

Abstract

Autophosphorylation of multifunctional Ca²⁺/calmod-ulin-dependent protein kinase converts it from a Ca²⁺-dependent to a Ca²⁺-independent or autonomous kinase, a process that may underlie some long-term enhancement of transient Ca²⁺ signals. We demonstrate that the neuronal α subunit clone expressed in COS-7 cells (α-CaM kinase) is sufficient to encode the regulatory phenomena characteristic of the multisubunit kinase isolated from brain. Activity of α-CaM kinase is highly dependent on Ca²⁺/calmodulin. It is converted by autophosphorylation to an enzyme capable of Ca²⁺-independent (autonomous) substrate phosphorylation and autophosphorylation. Using site-directed mutagenesis, we separately eliminate five putative auto-phosphorylation sites within the regulatory domain and directly examine their individual roles. Ca²⁺/calmodulin-dependent kinase activity is fully retained by each mutant, but Thr²⁸⁶ is unique among the sites in being indispensable for generation of an autonomous kinase.

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Neuron

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Article
Expression of a multifunctional Ca²⁺/calmodulin-dependent protein kinase and mutational analysis of its autoregulation

Deduced primary structure of the β subunit of brain type II Ca²⁺/calmodulin-dependent protein kinase determined by molecular cloning