| Summary: | 碩士 === 大同大學 === 生物工程學系(所) === 95 === In recent years, there has been an increasing attention on the cardiovascular health. Nattokinase, a highly fibrinolytic enzyme, has become the focus of study. In this study, the Vitreoscilla hemoglobin (VHb) gene was inserted into a previously constructed nattokinase gene- containing E. coli recombinant plasmid. The expression of VHb was intended to help the growth of bacteria and promote the expression level of target protein. The expression temperature and bacterial cell concentration at induction was adjusted to further facilitate the enzyme expression. With sorbitol and betaine-containing terrificbroth as the culture medium, the bacterial culture was induced after OD600=1.8, and the expression temperature was shifted from 37oC to 20 oC. The maximal intracellular nattokinase activity obtained was 8.7 U/ml, which occurred 8 hours after induction. The specific activity of purified nattokinase was 406.4 U/mg. The optimal temperature for nattokinase activity and Tm were 50 and 47℃, respectively. The optimal pH for nattokinase activity was 10, while the enzyme was most stable at pH 9.
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