Functional Expression and Characterization of Dipeptidyl Peptidase IV from the Black-Bellied Hornet Vespa basalis in Sf21 Insect Cells

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  • HSIEH Sheng-Kuo
    Graduate Institute of Biotechnology, National Chung-Hsing University
  • TZEN Jason T. C.
    Graduate Institute of Biotechnology, National Chung-Hsing University School of Chinese Medicine, China Medical University Agricultural Biotechnology Research Center, Academia Sinica
  • WU Tzong-Yuan
    Department of Bioscience Technology, Chung Yuan Christian University
  • CHEN Ying-Ju
    Department of Bioscience Technology, Chung Yuan Christian University
  • YANG Wei-Hung
    School of Chinese Medicine, China Medical University
  • HUANG Chun-Fa
    School of Chinese Medicine, China Medical University
  • HSIEH Feng-Chia
    Division of Biopesticides, Agricultural Chemicals and Toxic Substances Research Institute
  • JINN Tzyy-Rong
    School of Chinese Medicine, China Medical University

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  • Functional Expression and Characterization of Dipeptidyl Peptidase IV from the Black-Bellied Hornet<i>Vespa basalis</i>in Sf21 Insect Cells

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The maturation of mastoparan B, the major toxin peptide in the venom of Vespa basalis, requires enzymatic cleavage of its prosequence presumably via sequential liberation of dipeptides. The putative processing enzyme, dipeptidyl peptidase IV, was expressed as a glycosylated His-tag fusion protein (rDPP-IV) via the baculovirus expression system. rDPP-IV purified by one-step nickel-affinity chromatography was verified by Western blot and LC-MS/MS analysis. The kcat/Km of rDPP-IV was determined to be in the range of 10–500 mM−1·S−1 for five synthetic substrates. The optimal temperature and pH for rDPP-IV were determined to be 50 °C and pH 9. Enzymatic activity of rDPP-IV was significantly reduced by 80 and 60% in the presence of sitagliptin and phenylmethylsulfonyl fluoride respectively.

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