A brain-specific Ca2+/calmodulin-dependent protein kinase (CaM kinase-Gr) is regulated by autophosphorylation. Relevance to neuronal Ca2+ signaling

M.V Frangakis,C.A Ohmstede,N Sahyoun

doi:10.1016/s0021-9258(18)99164-7

M.V Frangakis, C.A Ohmstede + Show 1 more

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https://doi.org/10.1016/s0021-9258(18)99164-7

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Abstract

A neuronal Ca2+/calmodulin-dependent protein kinase (CaM kinase-Gr) undergoes autophosphorylation on a serine residue(s) in response to Ca2+ and calmodulin. Phosphate incorporation leads to the formation of a Ca(2+)-independent (autonomous) activity state, as well as potentiation of the Ca2+/calmodulin-dependent response. The autonomous enzyme activity of the phosphorylated enzyme approximately equals the Ca2+/calmodulin-stimulated activity of the unphosphorylated enzyme, but displays diminished affinity toward ATP and the synthetic substrate, syntide-2. The Km(app) for ATP and syntide-2 increased 4.3- and 1.7-fold, respectively. Further activation of the autonomous enzyme by Ca2+/calmodulin yields a marked increase in the affinity for ATP and peptide substrate such that the Km(app) for ATP and syntide-2 decreased by 14- and 8-fold, respectively. Both autophosphorylation and the addition of Ca2+/calmodulin are required to produce the maximum level of enzyme activation and to increase substrate affinity. Unlike Ca2+/calmodulin-dependent protein kinase type II that is dephosphorylated by the Mg(2+)-independent phosphoprotein phosphatases 1 and 2A, CaM kinase-Gr is dephosphorylated by a Mg(2+)-dependent phosphoprotein phosphatase that may be related to the type 2C enzyme. Dephosphorylation of CaM kinase-Gr reverses the effects of autophosphorylation on enzyme activity. A comparison between the autophosphorylation and dephosphorylation reactions of CaM kinase-Gr and Ca2+/calmodulin-dependent protein kinase type II provides useful insights into the operation of Ca(2+)-sensitive molecular switches.

Highlights

A neuronal Ca2+/calmodulin-dependentprotein ki- function is unknown, represents a portion of CaM kinase-Gr nase (CaM kinase-Gr) undergoes autophosphorylation that contains the calmodulin-binding and polyglutamate-rich on a serine residue(s) inresponse to Ca2+and calmod- regions but lacks a protein kinase catalytic domain[4]
Phosphate incorporation leads to the formation of CaMkinase-Gr differs from amajor brain calmodulina Ca2+-independen(tautonomous)activity state, as welldependent kinase, CaM kinase-11, in several aspects such as as potentiation of the Ca’+/calmodulin-dependentre- amino acid sequence, subcellular distribution, subunitmolecsponse
Unlike Ca2+/calmodulin-dependentprotein kinase Both polypeptide components (Mr 65,000 and 67,000) of type I1 that is dephosphorylatedby the Mg2+-independ- CaM kinase-Gr readily undergo autophosphorylation in the ent phosphoprotein phosphatases 1 and 2A, CaM ki- presence of Ca2+ andcalmodulin [1].Autophosphorylation of nase-Gr is dephosphorylated by a Mg2+-dependent CaM kinase-I1 has been examined intensively and found to phosphoproteinphosphatase that may berelated to the type 2C enzyme

Summary

Introduction

A neuronal Ca2+/calmodulin-dependentprotein ki- function is unknown, represents a portion of CaM kinase-Gr nase (CaM kinase-Gr) undergoes autophosphorylation that contains the calmodulin-binding and polyglutamate-rich on a serine residue(s) inresponse to Ca2+and calmod- regions but lacks a protein kinase catalytic domain[4]. Aliquots containing 0.2 pmol of protein kinase were removed and immediately assayedfor syntide-2 phosphorylation enzyme thatarebroughtabout by theaddition of Ca2+- for 6 min a t 23 "C in a reaction mixture containing 60 p M syntide-2, calmodulin.

Results

Conclusion