Expression, purification and kinase activity analysis of maize ZmSPK1, a member of plant SnRK2 subfamily

Abstract

Kinase activity is essential for a protein kinase to perform its biological function. In previous study we have cloned a novel plant SnRK2 subfamily gene from maize and named it as ZmSPK1. In this study the
cDNA of ZmSPK1 with dHA-His6 tag was amplified by PCR and was subcloned into the yeast expression vector p426GAL1. The constructed plasmid named as p426GAL1-SPK1-dHA-His6 was transformed into Saccharomyces cerevisiae. Under the induction of galactose the recombinant protein was expressed. The purified recombinant protein showed a single specific band by the analysis of the western-blot. In an in vitro kinase assay, the purified recombinant protein showed phosphorylation and autophosphorylation activity. This showed that ZmSPK1 encodes a functional protein kinase.