Autophosphorylation-Dependent Activation of a Calcium-Dependent Protein Kinase from Groundnut1
AUTOR(ES)
Chaudhuri, Subho
FONTE
American Society of Plant Physiologists
RESUMO
Ca2+-dependent protein kinases (CDPKs) containing a calmodulin-like domain integrated in their primary sequence are present primarily in plants. A member of this family was characterized from the groundnut (Arachis hypogea) plant and called GnCDPK (M. DasGupta [1994] Plant Physiol 104: 961–969). GnCDPK specifically uses the myosin light chain synthetic peptide (MLCpep), which is the phosphate-accepting domain of smooth muscle myosin light chains (KKRPQRATSNVFS), as an exogenous substrate under in vitro experimental conditions. In this report we show that GnCDPK undergoes intramolecular autophosphorylation. This self-phosphorylation occurs in threonine residues in a Ca2+-dependent (K0.5 = 0.5 μm) and calmodulin-independent manner. The kinase activity toward MLCpep and its sensitivity to Ca2+ were unaffected by prior autophosphorylation when measured under saturating ATP concentrations. The role of autophosphorylation in the exogenous substrate MLCpep phosphorylation reaction was reinvestigated at low ATP concentrations. A pronounced lag time of 1 to 2 min, followed by a linear increase of activity for 7.5 min, was seen in the initial rate of MLCpep phosphorylation under such suboptimal conditions. Prior autophosphorylation completely abolished this lag phase, and a sharp rise of exogenous substrate phosphorylation was seen from the 1st min. Our results suggest that autophosphorylation is a prerequisite for the activation of GnCDPK.
ACESSO AO ARTIGO
http://www.pubmedcentral.nih.gov/articlerender.fcgi?artid=59325Documentos Relacionados
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