Differential inhibition of plant calcium-dependent protein kinases by long chain fatty acids and other amphiphiles

Abstract

Ca 2+-dependent protein kinase (CDPK) was partially purified from wheat ( Triticum aestivum) embryo and from wheat and silver beet ( Beta vulgaris) leaves by extensive protocols involving Ca 2+-dependent hydrophobic chromatography on phenyl-Sepharose CL-4B, ion exchange chromatography on DEAE-Sephacel and gel filtration as common procedures. Wheat leaf CDPK is similar to wheat embryo CDPK (purified by application of the same protocol) in Ca 2+-dependence, substrate specificity, molecular size (mol. wt. 81 000) and in inhibition by the calmodulin antagonist trifluoperazine. Both wheat leaf and wheat embryo CDPKs phosphorylate histones (III-S and II-AS preparations), bovine serum albumin, casein and myosin light chains but not the synthetic cyclic AMP-dependent protein kinase substrate, Kemptide. Wheat and silver beet CDPKs are inhibited by amphipathic ligands including even carbon number long chain fatty acids (C 14C 22), odd carbon number long fatty acids (C 15C 21), long chain aliphatic alcohols (C 8C 11), amino acridines and sphingosine. Behenic acid and arachidic acid are potent inhibitors of wheat embryo CDKP (IC 50 values 50 and 60 μM, respectively) but are poor inhibitors of the wheat leaf and silver beet leaf CDPKs.