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Literature summary extracted from

  • Miller, J.B.; Pratap, A.; Miyahara, A.; Zhou, L.; Bornemann, S.; Morris, R.J.; Oldroyd, G.E.
    Calcium/calmodulin-dependent protein kinase is negatively and positively regulated by calcium, providing a mechanism for decoding calcium responses during symbiosis signaling (2013), Plant Cell, 25, 5053-5066.
    View publication on PubMedView publication on EuropePMC

Activating Compound

EC Number Activating Compound Comment Organism Structure
2.7.11.17 additional information the enzyme is activated by calmdulin binding at higher Ca2+ concentrations, autoactivation of CCaMK by mutation of Thr271 Medicago truncatula

Cloned(Commentary)

EC Number Cloned (Comment) Organism
2.7.11.17 recombinant expression of MBP-fusion wild-type enzyme and of His-tagged mutant enzymes in Escherichia coli strain BL21 (DE3), expression of recombinant GST-tagged wild-type enzyme in Escherichia coli strain Rosetta (DE3), recombinant expression of the kinase domain of CCaMK alone gives rise to spontaneous nodulation 8 weeks after root transformation, but in the absence of rhizobia Medicago truncatula

Protein Variants

EC Number Protein Variants Comment Organism
2.7.11.17 E319A site-directed mutagenesis, Thr271 phosphorylation is reduced in the mutant compared to the wild-type Medicago truncatula
2.7.11.17 L324A site-directed mutagenesis, Thr271 phosphorylation is abolished in the L324A mutant compared to the wild-type Medicago truncatula
2.7.11.17 L333A site-directed mutagenesis, Thr271 phosphorylation is enhanced in the L324A mutant compared to the wild-type, resulting in a constitutively active enzyme Medicago truncatula
2.7.11.17 R323A site-directed mutagenesis, the mutant is unable to autoactivate Medicago truncatula
2.7.11.17 S322A site-directed mutagenesis, the mutant is able to autoactivate Medicago truncatula
2.7.11.17 S343A site-directed mutagenesis, Thr271 phosphorylation is reduced in the mutant compared to the wild-type Medicago truncatula
2.7.11.17 T271A site-directed mutagenesis, the mutant is able to autoactivate Medicago truncatula
2.7.11.17 T271D site-directed mutagenesis, the mutant is able to autoactivate Medicago truncatula
2.7.11.17 T271I site-directed mutagenesis, the mutant is able to autoactivate Medicago truncatula
2.7.11.17 T271X autoactivation of CCaMK by mutation of Thr271 Medicago truncatula

Metals/Ions

EC Number Metals/Ions Comment Organism Structure
2.7.11.17 Ca2+ required for binding of cofactor calmodulin, differential calcium binding affinities of the EF-hand domains of the enzyme. Ca2+ binding via the EF-hand domains promotes Thr271 phosphorylation and enzyme deactivation, a deactivating conformation of CCaMK is stabilized by Thr271 phosphorylation. The binding of Ca2+ negatively regulates CCaMK by enhancing Thr271 phosphorylation Medicago truncatula
2.7.11.17 Mg2+ required Medicago truncatula

Organism

EC Number Organism UniProt Comment Textmining
2.7.11.17 Medicago truncatula Q6RET7 cv. Jemalong A17
-

Posttranslational Modification

EC Number Posttranslational Modification Comment Organism
2.7.11.17 phosphoprotein enzyme phosphorylation at residue Thr271, CCaMK autophosphorylation modeling, overview Medicago truncatula

Purification (Commentary)

EC Number Purification (Comment) Organism
2.7.11.17 recombinant tagged wild-type enzymes from Escherichia coli strains by affinity chromatography, recombinant His-tagged mutant enzymes from Escherichia coli strain BL21 (DE3) by nickel affinity chromatography and gel filtration Medicago truncatula

Subunits

EC Number Subunits Comment Organism
2.7.11.17 More modelling of two stable states for CCaMK: an inactive form that predominates at basal Ca2+ concentrations and involves a hydrogen bond network between the CaM binding and kinase domains and an active form that predominates during Ca2+ oscillations associated with CaM binding Medicago truncatula

Synonyms

EC Number Synonyms Comment Organism
2.7.11.17 calcium/calmodulin-dependent protein kinase
-
 Medicago truncatula
2.7.11.17 CCaMK
-
 Medicago truncatula
2.7.11.17 ccamk-1
-
 Medicago truncatula
2.7.11.17 DMI-3
-
 Medicago truncatula
2.7.11.17 dmi3-1
-
 Medicago truncatula
2.7.11.17 TRV25
-
 Medicago truncatula

Temperature Optimum [°C]

EC Number Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
2.7.11.17 30
-
 assay at Medicago truncatula

pH Optimum

EC Number pH Optimum Minimum pH Optimum Maximum Comment Organism
2.7.11.17 7.5
-
 assay at Medicago truncatula

Cofactor

EC Number Cofactor Comment Organism Structure
2.7.11.17 ATP
-
 Medicago truncatula
2.7.11.17 Calmodulin required, calcium-dependent CaM binding overrides the effects of autophosphorylation and activates the protein Medicago truncatula

General Information

EC Number General Information Comment Organism
2.7.11.17 malfunction autoactivation of CCaMK by mutation of Thr271. The complete absence of the EF-hand domains leads to unregulated Thr271 phosphorylation and deactivation of the protein Medicago truncatula
2.7.11.17 additional information modelling of two stable states for CCaMK: an inactive form that predominates at basal Ca2+ concentrations and involves a hydrogen bond network between the CaM binding and kinase domains and an active form that predominates during Ca2+ oscillations associated with CaM binding. The hydrogen bond between phosphorylated Thr271 and Ser322 is the predominant stabilizing bond at the inactive fold in Medicago truncatula CCaMK Medicago truncatula
2.7.11.17 physiological function the differential calcium binding affinities of the EF-hand domains compared with those of calmodulin suggest that enzyme CCaMK is maintained in the inactive state at basal calcium concentrations and is activated via calmodulin binding during calcium oscillations. Recombinant expression of the kinase domain of CCaMK alone gives rise to spontaneous nodulation 8 weeks after root transformation, but in the absence of rhizobia. The binding of Ca2+ negatively regulates CCaMK by enhancing Thr271 phosphorylation, while binding of calmodulin activates it, the enzyme remains inactive at basal Ca2+ concentrations and becomes activated during Ca2+ spiking. Enzyme regulation modelling, overview Medicago truncatula