Characterization of Recombinant Human Chymase Expressed in Escherichia coli

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Author(s)

Abstract

We compared recombinant human chymase expressed in Escherichia coli with human chymase purified from vascular tissues.The recombinant chymase, the structure of which was NH<SUB>2</SUB>-enterokinase cleavage site-chymase-COOH, was expressed in Escherichia coli and then was solubilized and renatured.The protein did not have a chymase activity, but gained this activity after the cleavage of the N-terminal site by enterokinase.The enzyme was purified by heparin affinity and gel filtration columns.The N-terminal sequence of the protein was identical to the sequence for human chymase.The molecular weights of the recombinant chymase and chymase purified from human vascular tissues were 26 and 30kDa, respectively, and the 4kDa difference was thought to be due to the presence or absence of glycan.The optimum pH of the recombinant enzyme activity was between 7.5 and 9.0.The activity of the recombinant enzyme was inhibited by chymostatin, soybean trypsin inhibitor and phenylmethylsulfonyl fluoride, but not by ethylenediaminetetraacetic acid and aprotinin.This enzyme cleaved specifically the Phe<SUP>8</SUP>-His<SUP>9</SUP> bond of angiotensin(Ang)I to form Ang II and that of big endothelin(ET)-1 to form ET-1-(1-31).These findings demonstrated that the enzymatic characteristics of the recombinant enzyme were identical to that of native human chymase.

Journal

  • The Japanese Journal of Pharmacology

    The Japanese Journal of Pharmacology 82(2), 144-149, 2000-02-01

    The Japanese Pharmacological Society

References:  34

Cited by:  1

Codes

  • NII Article ID (NAID)
    10008183157
  • NII NACSIS-CAT ID (NCID)
    AA00691188
  • Text Lang
    ENG
  • Article Type
    Journal Article
  • ISSN
    00215198
  • NDL Article ID
    5283952
  • NDL Source Classification
    ZS51(科学技術--薬学)
  • NDL Call No.
    Z53-D199
  • Data Source
    CJP  CJPref  NDL  J-STAGE 
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