Isolation and Characterization of Recombinant Vibrio parahaemolyticus Collagenase

재조합 Vibrio parahaemolyticus 콜라겐분해효소의 분리 및 특성 분석

  • 차재호 (부산대학교 자연과학대학 미생물학과) ;
  • 김수광 (부산대학교 자연과학대학 미생물학과) ;
  • 전인준 (부산대학교 자연과학대학 미생물학과) ;
  • 이재원 (부산대학교 자연과학대학 미생물학과)
  • Published : 2003.06.01


The collagenase gene from Vibrio parahaemolyticus 04 was subcloned into an expression vector pET-29b. The recombinant collagenase was expressed in Escherichia coli BL2l(DE3) and partially purified by Hi-Trap affinity and Sephacryl S-100 size exclusion chromatographies. The recombinant enzyme was purified by 43.7-fold and the yield was 73%. SDS-PAGE revealed that the molecular weight of the enzyme was approximately 35 kDa. Substrate specificity study of the enzyme displayed that the enzyme showed the highest activity with the type I collagen and the synthetic peptide, Z-GPGGPA, indicating that the enzyme was indeed a collagenase. The enzyme showed broad pH optimum around pH 6-12 and was stable between pH 5.5 and 11.5. The optimum temperature for the type I collagen degradation was $35^{\circ}C$. The thermostability measurement of the enzyme indicated that the enzyme was stable up to $55^{\circ}C$, but the activity was diminished quickly above $60^{\circ}C.$


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