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The journal of microbiology v.50 no.2, 2012년, pp.332 - 340   SCIE SCOPUS
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Heterologous Expression of Polygalacturonase Genes Isolated from Galactomyces citri-aurantii IJ-1 in Pichia pastoris

Cho, Il-Jae    (Department of Biotechnology (BK21 Program), Chung-Ang University   ); Yeo, In-Cheol    (Department of Biotechnology (BK21 Program), Chung-Ang University   ); Lee, Nam-Keun    (Department of Biotechnology (BK21 Program), Chung-Ang University   ); Jung, Suk-Hee    (Department of Biotechnology (BK21 Program), Chung-Ang University   ); Hahm, Young-Tae    (Department of Biotechnology (BK21 Program), Chung-Ang University  );
  • 초록

    The objective of this work was to isolate the polygalacturonase genes of Galactomyces citri-aurantii IJ-1 harvested from rotten citrus peels and to heterologously express these genes in Pichia pastoris. Two polygalacturonase (PG) genes from G. citri-aurantii IJ-1 were obtained and tentatively named PG1 and PG2. The genes were cloned into $pPICZ{\alpha}C$ , and expressed in Pichia pastoris strain GS115 with a native signal peptide or the ${\alpha}$ -factor secretion signal peptide of Saccharomyces cerevisiae. All of the recombinant proteins were successfully secreted into the culture media and confirmed as a single band with a molecular weight of 35 to 38 kDa by SDS-PAGE. The specific enzyme activities of recombinant PG1 and PG2 purified by His-tag affinity resin were 4,749 and 6,719 U/mg, respectively, with an optimal pH and temperature of pH 4.0 and $50^{\circ}C$ . The Michaelis-Menten kinetic constants for PG1 and PG2, $K_m$ , were confirmed to be 0.94 and 0.84 mM, respectively. In the presence of $Mn^{2+}$ , the activity of PG1 and PG2 were increased to 160.8 and 146.4% of normal levels, respectively. In contrast, $Cu^{2+}$ and $Fe^{3+}$ acted as strong inhibitors to the PGs.


  • 주제어

    polygalacturonase .   Galactomyces citri-aurantii .   Pichia pastoris .   heterologous expression.  

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