Purification and Characterization of Pectate Lyase I and II from Fungus Stereum purpureum.

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  • Miyairi Kazuo
    Department of Biochemistry and Biotechnology, Faculty of Agriculture and Life Science, Hirosaki University
  • Nishida Kaori
    Department of Biochemistry and Biotechnology, Faculty of Agriculture and Life Science, Hirosaki University
  • Takarae Masaru
    Department of Biochemistry and Biotechnology, Faculty of Agriculture and Life Science, Hirosaki University
  • Shikanai Yumiko
    Department of Biochemistry and Biotechnology, Faculty of Agriculture and Life Science, Hirosaki University
  • Okuno Toshikatsu
    Department of Biochemistry and Biotechnology, Faculty of Agriculture and Life Science, Hirosaki University

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Other Title
  • 銀葉病菌のペクチン酸リアーゼI とII の精製と性質
  • Purification and Characterization of Pectate Lyase 1 and 2 from Fungus Stereum purpureum

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Abstract

Two pectate lyases (PLs I and II) were isolated from the culture filtrate of Stereum purpureum, the causative fungus of apple silver-leaf disease. The lyases were isolated using DE52 column chromatography, and their molecular masses were estimated to be 48 kDa by SDS-PAGE. After treatment with endo-β-N-acetylglucosaminidase (Endo H), their molecular masses decreased equally to 38 kDa, indicating that these PLs are glycoproteins. The pI values for PLs I and II were determined to be 4.15 and 4.10, respectively. The amino acid compositions of PLs I and II were very similar, and furthermore the amino acid sequences of the N-terminal 20 residues of the two enzymes were identical. Their optimum pH values, and thermal and pH stabilities were 9.5, up to 50°C, and pH 6-7, respectively. Both PLs I and II required 0.2 mM Ca2+ ion for maximum activity. The enzymes were assigned as endo-types because of the detection of oligo-GalUAs as products from the initial reaction. However they mainly formed products less than octa-GaIUA in degree of polymerization. Erwinia carotovora PL mainly formed products more than octa-GalUA. Therefore they were different in their initial reaction products. As in the case of the E. carotovora PL, the final products of S. purpureum PL were 4, 5-unsaturated di- and tri-GalUA.

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