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Improved isolation, stability and substrate specificity of cucumisin, a plant serine endopeptidase.

Authors
  • Kaneda, M
  • Yonezawa, H
  • Uchikoba, T
Type
Published Article
Journal
Biotechnology and applied biochemistry
Publication Date
Oct 01, 1995
Volume
22 ( Pt 2)
Pages
215–222
Identifiers
PMID: 7576259
Source
Medline
License
Unknown

Abstract

Cucumisin (EC 3.4.21.25), a serine endopeptidase, was isolated by a simple purification procedure from the prince melon (Cucumis melo ssp. melo, cv. 'Prince Melon'). The enzyme is stable over a wide pH range (4-11) and to heat, 80% of its initial activity remaining even at pH 11.1 and at 60 degrees C for 20 min. The enzyme was inactive at 72 degrees C and pH 8.0, but 38% of the activity remained in the presence of 10% (w/v) glycerol. Caseinolysis by cucumisin indicated full activity in 8 M urea at pH 9.1 and 50 degrees C. Cucumisin was inactivated by treatment with trypsin at 37 degrees C for 24 h, but was not affected by alpha-chymotrypsin. The synthetic substrates benzyloxycarbonyltyrosine nitrophenyl ester (Z-Tyr-ONp) and benzoyltyrosine ethyl ester (Bz-Tyr-OEt) were cleaved, but Z-Lys-ONp and tosylarginine methyl ester (Tos-Arg-OMe) were not cleaved by cucumisin. Oxidized insulin B-chain was hydrolysed by cucumisin at 37 degrees C for 24 h, 21 cleavage sites being detected. Cucumisin could not cleave the C-termini of all the valine residues in the oxidized insulin B-chain molecule.

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