Molecular Cloning and Characterization of Prolyl Endopeptidase from Human T Cells

Abstract

The prolyl endopeptidase (PEP) gene of human T cells was amplified by the PCR method and cloned in Escherichia coli. The complete gene consisted of 2, 130 nucleotides corresponding to 710 amino acid residues with a calculated molecular mass of 80, 750. The nucleotide sequence of this clone revealed that T cell PEP DNA is 48, 50, and 91% homologous to those of Flavobacterium meningosepticum, Aeromonas hydrophila, and porcine brain PEP, respectively. This gene was fused to the lacZ sequence from E. coli and expressed as a fused protein in E. coli. This fused protein exhibited PEP activity, which was inhibited by Z-Pro-prolinal, a specific inhibitor of PEP. The fused protein was purified on a β-galactosidase specific affinity column. A polyclonal antibody was raised against the purified protein. Immunological characterization suggested that this protein is different from cytosol-soluble PEP.