Abstract
Phospholipids, the major structural components of membranes, can also have functions in regulating signaling pathways in plants under biotic and abiotic stress. The effects of adding phospholipids on the activity of stress-induced calcium dependent protein kinase (CaCDPK1) from chickpea are reported here. Both autophosphorylation as well as phosphorylation of the added substrate were enhanced specifically by phosphatidylcholine and to a lesser extent by phosphatidic acid, but not by phosphatidylethanolamine. Diacylgylerol, the neutral lipid known to activate mammalian PKC, stimulated CaCDPK1 but at higher concentrations. Increase in Vmax of the enzyme activity by these phospholipids significantly decreased the Km indicating that phospholipids enhance the affinity towards its substrate. In the absence of calcium, addition of phospholipids had no effect on the negligible activity of the enzyme. Intrinsic fluorescence intensity of the CaCDPK1 protein was quenched on adding PA and PC. Higher binding affinity was found with PC (K½ = 114 nM) compared to PA (K½ = 335 nM). We also found that the concentration of PA increased in chickpea plants under salt stress. The stimulation by PA and PC suggests regulation of CaCDPK1 by these phospholipids during stress response.
Highlights
Plants have a novel family of protein kinases known as calcium dependent protein kinase
Calcium Dependent Protein Kinase 1 (CaCDPK1) was measured in presence or absence of crude phospholipids
Parallel gels were run with CaCDPK1 and histone, and stained with coomassie brilliant blue (CCB) for a loading control
Summary
Plants have a novel family of protein kinases known as calcium dependent protein kinase CDPKs are reported in Plasmodium [2], Paramecium [3], Taxoplasma [4]. They are not present in the eukaryotic genome of yeast (Saccharomyces cerevisiae), nematodes [5], fruit flies (Drosophila melanogaster) [6] and humans [7]. Many CDPKs are predicted to have myristoylation and palmitoylation sites at their N-terminal domain which are essential for membrane anchorage [25,26]
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