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http://dx.doi.org/10.5352/JLS.2020.30.2.137

Partial Purification of OsCPK11 from Rice Seedlings and Its Biochemical Characterization  

Shin, Jae-Hwa (Gyeonggi Science High School)
Kim, Sung-Ha (Department of Biology Education, Korea National University of Education)
Publication Information
Journal of Life Science / v.30, no.2, 2020 , pp. 137-146 More about this Journal
Abstract
Calcium is one of the important secondary signaling molecules in plant cells. Calcium-dependent protein kinases (CDPK)-the sensor proteins of Ca2+ and phosphorylating enzymes-are the most abundant serine/threonine kinases in plant cells. They convert and transmit signals in response to various stimuli, resulting in specific responses in plants. In rice, 31 CDPK gene families have been identified, which are mainly involved in plant growth and development and are known to play roles in response to various stress conditions. However, little is known about the biochemical characteristics of CDPK proteins. In this study, OsCPK11-a CDPK in rice-was partially purified, and its biochemical characteristics were found. Partially purified OsCPK11 from rice seedlings was obtained by three-step column chromatography that involved anion exchange chromatography consisting of DEAE, hydrophobic interaction chromatography consisting of phenyl-Sepharose, and gel filtration chromatography consisting of Sephacryl-200HR. An in vitro kinase assay using partially purified OsCPK11 was also performed. This partially purified OsCPK11 had a molecular weight of 54 kDa and showed a strong hydrophobic interaction with the hydrophobic resin. In vitro kinase assay showed that the OsCPK11 also had Ca2+-dependent autophosphorylation activity. The OsCPK11 phosphorylated histone III-S, and the optimum pH for its kinase activity was found to be 7.5~8.0. The native OsCPK11 shared several biochemical characteristics with recombinant OsCPK11 studied previously, and both had Ca2+-dependent autophosphorylation activity and favored histone III-S as a substrate for kinase activity, which also had a Ca2+-dependence.
Keywords
Autophosphorylation; $Ca^{2+}$-mediated signaling; CDPK; column chromatography; OsCPK11; rice; transphosphorylation;
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