Biochemical Characterization of An Arginine-Specific Alkaline Trypsin from Bacillus licheniformis |
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Authors: | Jin-Song Gong Wei Li Dan-Dan Zhang Min-Feng Xie Biao Yang Rong-Xian Zhang Heng Li Zhen-Ming Lu Zheng-Hong Xu Jin-Song Shi |
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Affiliation: | School of Pharmaceutical Science, Jiangnan University, Wuxi 214122, China; (J.-S.G.); (W.L.); (D.-D.Z.); (M.-F.X.); (B.Y.); (R.-X.Z.); (H.L.); (Z.-M.L.); (Z.-H.X.) |
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Abstract: | In the present study, we isolated a trypsin-producing strain DMN6 from the leather waste and identified it as Bacillus licheniformis through a two-step screening strategy. The trypsin activity was increased up to 140 from 20 U/mL through culture optimization. The enzyme was purified to electrophoretic homogeneity with a molecular mass of 44 kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and the specific activity of purified enzyme is 350 U/mg with Nα-Benzoyl-l-arginine ethylester as the substrate. The optimum temperature and pH for the trypsin are 65 °C and pH 9.0, respectively. Also, the enzyme can be significantly activated by Ba2+. This enzyme is relatively stable in alkaline environment and displays excellent activity at low temperatures. It could retain over 95% of enzyme activity after 180 min of incubation at 45 °C. The distinguished activity under low temperature and prominent stability enhance its catalytic potential. In the current work, the open reading frame was obtained with a length of 1371 nucleotides that encoded a protein of 456 amino acids. These data would warrant the B. licheniformis trypsin as a promising candidate for catalytic application in collagen preparation and leather bating through further protein engineering. |
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Keywords: | trypsin Bacillus licheniformis enzymatic properties cloning biocatalysis |
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