Dual regulation of a chimeric plant serine/threonine kinase by calcium and calcium/calmodulin

D Takezawa; S Ramachandiran; V Paranjape; B W Poovaiah

doi:10.1074/jbc.271.14.8126

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Dual regulation of a chimeric plant serine/threonine kinase by calcium and calcium/calmodulin

Journal article

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Dual regulation of a chimeric plant serine/threonine kinase by calcium and calcium/calmodulin

D Takezawa, S Ramachandiran, V Paranjape and B W Poovaiah

The Journal of biological chemistry, Vol.271(14), pp.8126-8132

04/05/1996

DOI: https://doi.org/10.1074/jbc.271.14.8126

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https://hdl.handle.net/2376/114568

PMID: 8626500

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Abstract

Amino Acid Sequence

Phosphorylation

Calmodulin - metabolism

Protein Structure, Secondary

Calcium - metabolism

Molecular Sequence Data

Structure-Activity Relationship

Recombinant Proteins

DNA Primers - chemistry

Recombinant Fusion Proteins - metabolism

Calcium-Calmodulin-Dependent Protein Kinases - chemistry

Base Sequence

Protein-Serine-Threonine Kinases - chemistry

Binding Sites

Calcium-Calmodulin-Dependent Protein Kinases - metabolism

A chimeric Ca2+/calmodulin-dependent protein kinase (CCaMK) gene characterized by a catalytic domain, a calmodulin-binding domain, and a neural visinin-like Ca2+-binding domain was recently cloned from plants (Patil, S., Takezawa, D., and Poovaiah, B. W. (1995) Proc. Natl. Acad. Sci. U. S. A. 92, 4797-4801). The Escherichia coli-expressed CCaMK phosphorylates various protein and peptide substrates in a Ca2+/calmodulin-dependent manner. The calmodulin-binding region of CCaMK has similarity to the calmodulin-binding region of the alpha-subunit of multifunctional Ca2+/calmodulin-dependent protein kinase (CaMKII). CCaMK exhibits basal autophosphorylation at the threonine residue(s) (0.098 mol of 32P/mol) that is stimulated 3.4-fold by Ca2+ (0.339 mol of 32P/mol), while calmodulin inhibits Ca2+-stimulated autophosphorylation to the basal level. A deletion mutant lacking the visinin-like domain did not show Ca2+-stimulated autophosphorylation activity but retained Ca2+/calmodulin-dependent protein kinase activity at a reduced level. Ca2+-dependent mobility shift assays using E. coli-expressed protein from residues 358 520 revealed that Ca2+ binds to the visinin-like domain. Studies with site-directed mutants of the visinin-like domain indicated that EF-hands II and III are crucial for Ca2+-induced conformational changes in the visinin-like domain. Autophosphorylation of CCaMK increases Ca2+/calmodulin-dependent protein kinase activity by about 5-fold, whereas it did not affect its Ca2+-independent activity. This report provides evidence for the existence of a protein kinase in plants that is modulated by Ca2+ and Ca2+/calmodulin. The presence of a visinin-like Ca2+-binding domain in CCaMK adds an additional Ca2+-sensing mechanism not previously known to exist in the Ca2+/calmodulin-mediated signaling cascade in plants.

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Title: Dual regulation of a chimeric plant serine/threonine kinase by calcium and calcium/calmodulin
Creators: D Takezawa - Department of Horticulture, Washington State University, Pullman, Washington 99164-6414, USA
S Ramachandiran
V Paranjape
B W Poovaiah
Publication Details: The Journal of biological chemistry, Vol.271(14), pp.8126-8132
Academic Unit: Horticulture, Department of
Publisher: United States
Identifiers: 99900547756301842
Language: English
Resource Type: Journal article