| Autor: |
Z.S. Xu, Y. Liang, J. Kong, S.S. Zhang, X.L. Liu, T. Wang |
| Jazyk: |
angličtina |
| Rok vydání: |
2022 |
| Předmět: |
|
| Zdroj: |
Journal of Dairy Science, Vol 105, Iss 7, Pp 5641-5653 (2022) |
| Druh dokumentu: |
article |
| ISSN: |
0022-0302 |
| DOI: |
10.3168/jds.2021-21699 |
| Popis: |
ABSTRACT: Streptococcus thermophilus is a common yogurt starter that consumes lactose as its primary carbon source. The enzyme β-galactosidase is essential for the lactose metabolism and the growth of this species. Streptococcus thermophilus appears to be a promising cell factory. Food-grade vectors have advantages in heterologous protein expression. This study aimed to determine whether the β-galactosidase of S. thermophilus has the α-complementary characteristic and to develop a novel food-grade vector based on this phenomenon. The N-terminal 7 to 36 AA residues of the β-galactosidase in S. thermophilus were deleted. The obtained mutant S. thermophilus Δα lost β-galactosidase activity and growth ability in the lactose medium. Subsequently, plasmids expressing α-fragments with different lengths of 1 to 36 (Sα1), 1 to 53 (Sα2), and 1 to 88 (Sα3) AA were constructed and transformed into S. thermophilus Δα. Recombinant S. thermophilus Δα expressing Sα2 or Sα3 recovered the ability to grow in the lactose medium, and their β-galactosidase activity accounted for 24.5% or 11.5% of the wild strain, respectively. These results indicated that the α-complementation system of β-galactosidase existed in S. thermophilus. Based on the characteristic, a food-grade vector pSEα was constructed. Except for Sα2, vector pSEα expressed the α-donor derived from E. coli β-galactosidase. This facilitated the construction of recombinant plasmids in E. coli DH5α and thus improved the transformation efficiency of S. thermophilus. Green fluorescent protein as a reporter protein could be highly expressed in S. thermophilus using this vector. As a result, pSEα is an efficient and safe vector for S. thermophilus with potential food applications. |
| Databáze: |
Directory of Open Access Journals |
| Externí odkaz: |
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