Regulation Mechanism Of Dual Specificity Protein Phosphatase 1 In Arabidopsis(AtDsPTP1)

The Mitogen Activated Protein Kinase(MAPK)signaling pathway is ubiquitous in all of the eukaryotes.When plants suffer from extracellular stimuli such as inflammation,cytokine induction,environmental stress etc,the kinases in MAPK cascades sequentially activate and finally participate in several important processes such as the growth and development in plants and the regulation of its immune defences.MKPs(MAPK phosphatases)is a family of protein phosphatase that specifically dephosphorylate MAPK and precisely regulate the intensity and persistence of MAPK signals.It has been reported that there are more than 20 MAPKs in Arabidopsis thaliana,whereas only 5 MKPs have been found.The relationship and regulation mechanisms between MAPKs and MKPs are still unclear.In the paper,we cloned the dual-specific protein phosphatase DsPTP1(dual specificity protein phosphatase 1)and its regulatory protein CaMs(Camodulins)from the cDNA library in Arabidopsis thaliana.We obtained high-concentration and high-purity recombinant proteins with a series of protein purification related techniques.We determined the activity of DsPTP1 to small molecule substrate pNPP by means of enzyme kinetics,and indicated that DsPTP1 is an active protein phosphatase.At the same time we also measured the activity of DsPTP1 in presence of CaM2,found that the binding of CaM2 could enhance the affinity of DsPTP1 to pNPP and increase the activity as result.In addition,we vertified that DsPTP1 could dephosphorylate its physiological substrate MPK4,and the binding of CaM2 would inhibit its catalytic ability with western blot.In order to explain this phenomenon,we try to understand the molecular mechanism between DsPTP1 and MPK4,regulatory mechanism between DsPTPl and different CaMs using structural technology.However,as far as the current researches are concerned,we just obtain the microcrystal of DsPTP1 and the crystal of the complex of DsPTP1 and CaM2 has not been screened out.