Characterization of a Panel of Strong Constitutive Promoters from Streptococcus thermophilus for Fine-Tuning Gene Expression.

Streptococcus thermophilus, one of the most important probiotic lactic acid bacteria (LAB), is widely used in the dairy industry and attracts a lot of attention in metabolic engineering and synthetic biology. However, the available well-characterized constitutive promoters are rather limited to modulate gene expression in S. thermophilus. Here, a pool of constitutive promoters was screened by RNA-seq analysis and characterized in S. thermophilus, Lactobacillius casei, and Escherichia coli using the reporter red fluorescent protein. To assess their application potential, six constitutive promoters were selected for the expression of superoxide dismutase and significantly improved enzyme activity in the above three bacteria. Moreover, two strong constitutive promoters were used to construct a dual-expression vector for overexpressing epsA and epsE, key proteins of exopolysaccharide (EPS) biosynthesis, resulting in the change of molecular weight and the titer of EPS. Taken together, this is the first well-characterized constitutive promoter library from S. thermophilus, which could be used as a basic toolbox for various applications in LAB and other bacteria.