Abstract
The xylose operon is an efficient biological element used for the regulation of gene expression in Bacillus licheniformis. Although the mechanism underlying the xylose-mediated regulation of this operon has been elucidated, the transcriptional changes that occur under various fermentation conditions remain unclear. In this study, the effects of different conditions on xylose operon expression were investigated. Significant upregulation was observed during the transition from the logarithmic phase to the stationary phase (2.5-fold, n = 3, p < 0.01). Glucose suppressed transcription over 168-fold (n = 3, p < 0.01). Meanwhile, the inhibitory effect of glucose hardly strengthened at concentrations from 20 to 180 g/L. Furthermore, the transcription of the xylose operon increased at elevated temperatures (25–42 °C) and was optimal at a neutral pH (pH 6.5–7.0). Based on these findings, relevant fermentation strategies (delaying the induction time, using dextrin as a carbon source, increasing the fermentation temperature, and maintaining a neutral pH) were proposed. Subsequently, these strategies were validated through the use of maltogenic amylase as a reporter protein, as an 8-fold (n = 3, p < 0.01) increase in recombinant enzyme activity compared to that under unoptimized conditions was observed. This work contributes to the development of fermentation optimization and furthers the use of the xylose operon as an efficient expression element.
Highlights
Bacillus licheniformis is a generally recognized as safe (GRAS) workhorse that has simple culture requirements and abundant protein secretion
Real-time quantitative PCR (RT-qPCR) was used to assess the transcriptional changes in the xylose operon in B. licheniformis at different growth stages, glucose stress levels, temperatures, and pH values
We explored the glucose-mediated carbon catabolite repression (CCR) on the xylose operon in B. licheniformis at glucose concentrations ranging from 20 to 200 g/L
Summary
Bacillus licheniformis is a generally recognized as safe (GRAS) workhorse that has simple culture requirements and abundant protein secretion. Previous studies have generally not investigated the dynamic transcriptional changes in the xylose operon during the fermentation process. As the function of an operon can be fine-tuned by growth stage, glucose stress level, temperature, or pH, the dynamic regulatory characteristics of an operon can indicate the generation of the product on the most essential level This fine tuning can serve as a reliable basis for process optimization [15]. RT-qPCR was used to assess the transcriptional changes in the xylose operon in B. licheniformis at different growth stages, glucose stress levels, temperatures, and pH values. The results revealed characteristics of the xylose operon with respect to the regulation of target gene expression during various fermentation processes and provided a scientific reference for fermentation optimization. A.n2T.s3rca.rniTpsrtcaironinpsatcilroiAnpatciltoiAnviac2tlty.iA3vo.icftTtytirhvoaeifntXytshcyoerlfioXptshteyieolOonXspayelelOroAospnceetOiruvonpintdeyreuoronnfGdtuelhrunecGdXoelsryuelcGoSosltseureecOSsostpsreeerSsostnreusns der Glucose Stress GluGcolsueGc,olasuesc,oaasesc,oaamscpoaemtciptoievmGteilptuiaevcnteoiatsilaevon,egaaulasoengaatuoloceogxtmuyoelpoxestyoetil,toxiisvyseel,opaisrsne,sapulirosmegspeuurdemestteuoodmbxteyeodlotbhsteoee,tbmhiseeopstmhtreeoisnsmuftlmuoinseetfndlutiinteaofnllutbfiaeecnltttofhiarecltmofarcotsotrinfluential factor rgigtnregirgitgnhegeritrnhegegutrhelaegtuiroelangtturiooilgfangtiheoorefnintxhogyfeltothxhsyeeelrooxespygeeluorolosapenteioro[op3nn0eo,r3[of31n0t]h.,3[eH310]xo.,y3wHl1oe]os.vweHereoo,vpwteehrere,ovtdnehree[,g3tdr0hee,e3g1dro]e.efegHinrooehfewiibnoeihvftiieoibnrni,htitciohboneintifcdoeoenrngrefcredoerenrefoedfrriendhibition conferred
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