Comparison of gal–lac operons in wild-type galactose-positive and -negative Streptococcus thermophilus by genomics and transcription analysis
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Streptococcus thermophilus is one of the most important homo-fermentative thermophilic bacteria, which is widely used as a starter culture in dairy industry. Both wild-type galactose-negative (Gal−) S. thermophilus AR333 and galactose-positive (Gal+) S. thermophilus S-3 in this study were isolated from Chinese traditional dairy products. Here, to access the mechanism of the difference of galactose utilization between strains AR333 and S-3, the expression of gal–lac operons was examined using real-time qPCR in the presence of different sugars, and the gene organization of gal–lac operons was characterized using comparative genomics analysis. As compared with medium containing glucose, the expression of gal–lac operons in AR333 and S-3 was significantly activated (> 5-fold) in the presence of galactose or lactose in the medium. More importantly, the expression of gal operon in S-3 was higher than that of AR333, suggesting that the strength of gal promoter in AR333 and S-3 may be different. The genomes of AR333 and S-3 were the first time sequenced to provide insight into the difference of gal–lac operons in these two strains. Comparative genomics analysis showed that gene order and individual gene size of gal–lac operons are conserved in AR333 and S-3. The DNA sequence of gal operon responsible for galactose utilization between AR333 and S-3 is almost identical except that galK promoter of S-3 possesses single base pair mutation (G to A substitution) at -9 box galK region. Moreover, the expression of red fluorescent protein can be activated by galK promoter of S-3, but cannot by galK promoter of AR333 in galactose medium, suggesting that gal operon is silent in AR333 and active in S-3 under galactose-containing medium. Overall, our results indicated that single point mutation at -9 box in the galK promoter can significantly affect the expression of gal operon and is largely responsible for the Gal+ phenotype of S. thermophilus.
Keywordsgal operon Galactose positive galK promoter Streptococcus thermophilus
This work was supported by the National Natural Science Foundation of China (Grant no. 31871776 and 31771956), National Key R&D Program of China (Grant no. 2018YFD0502306), Natural Science Foundation of Shanghai (Grant no. 18ZR1426800), Guangdong Sci. & Tech project (No. 2014B020201001 and 2017A030313149), Nansha District Sci. & Tech project (2015GG003), and “Shuguang Program” by Shanghai Education Development Foundation and Shanghai Municipal Education Commission (No. 15SG42).
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