【摘 要】
：
PTPMEG1 is an intracellular protein tyrosine phosphatase(PTP), which contains FERM and PDZ domains. This study focuses our attention on the expression, purifica
【机 构】
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Clinical Laboratory,Edmond H. Fischer Signal Transduction Laboratory,Key Laboratory for Molecular En
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PTPMEG1 is an intracellular protein tyrosine phosphatase(PTP), which contains FERM and PDZ domains. This study focuses our attention on the expression, purification and characterization of catalytic domain of PTPMEG1((MEG1) and preparation of its polyclonal antibody. A cDNA fragment encoding (MEG1 protein(amino acid residues 643―926) was amplified by PCR and then cloned into the pT7-7 vector. Both soluble and insoluble recombinant (MEG1 proteins were observed after induction by IPTG. Soluble (MEG1 was purified via two chromatographic steps, and the purified enzyme was characterized. With para-nitrophenylphosphate(pNPP) as a substrate, (MEG1 exhibited typical enzymatic characteristics of classic PTPs and classical Michaelis-Menten kinetics. Insoluble (MEG1, which was mainly distributed in the inclusion body of E. coli cells extracts, was purified by preparative electrophoresis gel for the preparation of the polyclonal antibody. A rabbit was immunized with (MEG1 purified by preparative electrophoresis to generate anti-(MEG1 antibody. Anti-serum was collected on 28th day after initial injection and purified via affinity chromatography. The purified polyconal antibody displayed a satisfactory titer and sensitivity.
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