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Papers In Press, published online ahead of print May 23, 2002
Department of Plant Sciences, University of Arizona, Tucson, AZ 85721
Corresponding Author: jkzhu{at}ag.arizona.edu
The Arabidopsis SOS2 (Salt Overly Sensitive 2)-like protein kinases (PKSes) are novel protein kinases that contain an SNF1-like catalytic domain with a putative activation loop and a regulatory domain with a FISL motif that binds calcium sensors. Very little biochemical and functional information is currently available on this family of kinases. Here, we report on the expression of the PKS11 gene, activation and characterization of the gene product, and transgenic evaluation of its function in plants. PKS11 transcript was preferentially expressed in roots of Arabidopsis plants. Recombinant glutathione-S-transferase (GST) fusion protein of PKS11 was inactive in substrate phosphorylation. However, the kinase can be highly activated by a threonine 161 to aspartate substitution (designated PKS11T161D) in the putative activation loop. Interestingly, PKS11 can also be activated by substitution of either a serine or tyrosine with aspartate within the activation loop. Deletion of the FISL motif also resulted in a slight activation of PKS11. PKS11T161D displayed an uncommon preference for Mn2+ over Mg2+ for substrate phosphorylation and autophosphorylation. The optimal pH and temperature values of PKS11T161D were determined to be 7.5 and 30 ºC, respectively. The activated kinase showed substrate specificity, and high affinity and catalytic efficiency for a peptide substrate p3 and for ATP. AMP or ADP at concentrations from 10 µM to 1 mM did not activate PKS11T161D. Transgenic Arabidopsis plants expressing PKS11T161D were more resistant to high concentrations of glucose, suggesting the involvement of this protein kinase in sugar signaling in plants. These results provide insights into the function as well as regulation and biochemical properties of the PKS protein kinase.
J. Biol. Chem, 10.1074/jbc.M107719200
Submitted on August 13, 2001
Revised on May 23, 2002
Accepted on May 23, 2002
Biochemical and functional characterization of PKS11, a novel Arabidopsis protein kinase
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