Structure Analysis, Prokaryotic Expression, and Subcellular Location of a New Protein Phosphatase 2C Protein from Maize

摘要

ZmPP2C (AY621066) is a protein phosphatase 2C 111KNA that we cloned from maize previously. TMpred program analysis indicated this protein contains a significant transmembrane helix of about 20 amino acid residues; however SVMtm predicted it is not a transmembrane protein. STRIDE, Modeller and RasMol program analysis suggested the ZmPP2C protein is a globular protein containing seven alpha helixes, fourteen beta sheets, twenty-two turns and one 310 helix. The coding region of ZmPP2C mRNA was sub-cloned into expression vectors, pET30a-c(+), and introduced into E. coli 151.21 (DE3) for expression. SDS-PAGE analysis indicated ZmPP2C was highly expressed at 37°C for 4 h with induction by 1 niM IPTG. Ultrasonic extraction experiment showed this recombinant protein was soluble in lysis buffer solution. The hexahistidine-tagged ZmPP2C fusion protein were purified and used to immunize rat for producing antibody. Protein gel blot analysis stated clearly that a specific polyclonal antibody against the protein was produced and can be used for further study about the ZmPP2C gene. Subcellular localization suggested that ZmPP2C protein was located in cell nucleus.