Biochemical characterization of a recombinant Swainsona canescens calcium-dependent protein kinase (ScCPK1)

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• 作者：S.M. Srideshikan ; V.L. Vanishree ; C. Jayabaskaran ; ^{cjb@biochem.iisc.ernet.in} ; ^{satpal_singh9@yahoo.com}
• 关键词：Calmidazolium ; Enzyme kinetics ; Phosphoamino acid ; Recombinant ScCPK1 ; Swainsona canescens
• 刊名：Plant Physiology and Biochemistry
• 出版年：2012
• 出版时间：May, 2012
• 年：2012
• 卷：54
• 期：Complete
• 页码：27-33
• 全文大小：592 K

文摘

Calcium-dependent protein kinases (CPKs) constitute a unique family of kinases involved in many physiological responses in plants. Biochemical and kinetic properties of a recombinant Swainsona canescens calcium-dependent protein kinase (ScCPK1) were examined in this study. The optimum pH and temperature for activity were pH 7.5 and 37?¡ãC, respectively. Substrate phosphorylation activity of ScCPK1 was calmodulin (CaM) independent. Yet CaM antagonists, W7 [N-(6-aminohexyl)-5-chloro-1-naphthalene sulphonamide] and calmidazolium inhibited the activity with IC₅₀ values of 750?nM and 350?¦ÌM, respectively. Both serine and threonine residues were found to be phosphorylated in autophosphorylated ScCPK1 and in histone III-S phosphorylated by ScCPK1. The [Ca²⁺] for half maximal activity (K_0.5) was found to be 0.4?¦ÌM for ScCPK1 with histone III-S as substrate. Kinetic analysis showed that K_M of ScCPK1 for histone III-S was 4.8?¦ÌM. These data suggest that ScCPK1 is a functional Ser/Thr kinase, regulated by calcium, and may have a role in Ca²⁺-mediated signaling in S.?canescens.