Journal of Microbiology and Biotechnology

  • 제17권4호
  • /
  • Pages.624-631
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  • 2007
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  • 1017-7825(pISSN)
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  • 1738-8872(eISSN)
한국미생물·생명공학회 (The Korean Society for Microbiology and Biotechnology)
권호 표지

Purification and Characterization of Two Thermostable Proteases from the Thermophilic Fungus Chaetomium thermophilum

  • Li, An-Na (Department of Environmental Biology, Shandong Agricultural University) ;
  • Ding, AI-Yun (Department of Environmental Biology, Shandong Agricultural University) ;
  • Chen, Jing (Department of Environmental Biology, Shandong Agricultural University) ;
  • Liu, Shou-An (Department of Environmental Biology, Shandong Agricultural University) ;
  • Zhang, Ming (Department of Environmental Biology, Shandong Agricultural University) ;
  • Li, Duo-Chuan (Department of Environmental Biology, Shandong Agricultural University)
  • 발행 : 2007.04.30
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초록

Thermostable protease is very effective to improve the industrial processes in many fields. Two thermostable extracellular proteases from the culture supernatant of the thermophilic fungus Chaetomium thermophilum were purified to homogeneity by tractional ammonium sulfate precipitation, ion-exchange chromatography on DEAE-Sepharose, and Phenyl-Sepharose hydrophobic interaction chromatography. By SDS-PAGE, the molecular mass of the two purified enzymes was estimated to be 33 kDa and 63 kDa, respectively. The two proteases were found to be inhibited by PMSF, but not by iodoacetamide and EDTA. The 33 kDa protease (PRO33) exhibited maximal activity at pH 10.0 and the 63kDa protease (PRO63) at pH5.0. The optimum temperature for the two proteases was $65^{\circ}C$. The PRO33 had a $K_m$ value of 6.6mM and a $V_{max}$ value of $10.31{\mu}mol/l/min$, and PRO63 l7.6mM and $9.08{\mu}mol/l/min$, with casein as substrate. They were thermostable at $60^{\circ}C$. The protease activity of PRO33 and PRO63 remained at 67.2% and 17.31%, respectively, after incubation at $70^{\circ}C$ for 1h. The thermal stability of the two enzymes was significantly enhanced by $Ca^{2+}$. The residual activity of PRO33 and PRO63 at $70^{\circ}C$ after 60min was approximately 88.59% and 39.2%, respectively, when kept in the buffer containing $Ca^{2+}$. These properties make them applicable for many biotechnological purposes.

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