Heterologous expression of Streptomyces coelicolor trehalose synthase and whole-cell biocatalyst production of trehalose in Escherichia coli.
- Author:
Ao WU
1
;
Xian ZHANG
1
;
Meijuan XU
1
;
Taowei YANG
1
;
Huazhong LI
1
;
Zhiming RAO
1
Author Information
- Publication Type:Journal Article
- Keywords: clone and expression; site-directed mutagenesis; trehalose synthase; whole-cell conversion
- MeSH: Biocatalysis; Cloning, Molecular; Escherichia coli; Glucosyltransferases; Streptomyces coelicolor; Trehalose
- From: Chinese Journal of Biotechnology 2019;35(7):1348-1358
- CountryChina
- Language:Chinese
- Abstract: The trehalose synthase (ScTreS) gene from Streptomyces coelicolor was successfully cloned and heterologously expressed in Escherichia coli BL21(DE3). The protein purified by Ni-NTA affinity column showed an apparent molecular weight (MW) of 62.3 kDa analyzed by SDS-PAGE. The optimum temperature of the enzyme was 35 °C and the optimum pH was 7.0; the enzyme was sensitive to acidic conditions. By homologous modeling and sequence alignment, the enzyme was modified by site-directed mutagenesis. The relative activities of the mutant enzymes K246A and A165T were 1.43 and 1.39 times that of the wild type, an increased conversion rate of 14% and 10% respectively. To optimize the synthesis conditions of trehalose, the mutant strain K246A was cultivated in a 5-L fermentor and used for whole-cell transformation. The results showed that with the substrate maltose concentration of 300 g/L at 35 °C and pH 7.0, the highest conversion rate reached 71.3%, and the yield of trehalose was 213.93 g/L. However, when maltose concentration was increased to 700 g/L, the yield of trehalose can reach 465.98 g/L with a conversion rate of 66%.
