Intelligent microbial cell factory with genetic pH shooting (GPS) for cell self-responsive base/acid regulation

Abstract

Background In industrial fermentation, pH fluctuation resulted from microbial metabolism influences the strain performance and the final production. The common way to control pH is adding acid or alkali after probe detection, which is not a fine-tuned method and often leads to increased costs and complex downstream processing. Here, we constructed an intelligent pH-sensing and controlling genetic circuits called “Genetic pH Shooting (GPS)” to realize microbial self-regulation of pH. Results In order to achieve the self-regulation of pH, GPS circuits consisting of pH-sensing promoters and acid-/alkali-producing genes were designed and constructed. Designed pH-sensing promoters in the GPS can respond to high or low pHs and generate acidic or alkaline substances, achieving endogenously self-responsive pH adjustments. Base shooting circuit (BSC) and acid shooting circuit (ASC) were constructed and enabled better cell growth under alkaline or acidic conditions, respectively. Furthermore, the genetic circuits including GPS, BSC and ASC were applied to lycopene production with a higher yield without an artificial pH regulation compared with the control under pH values ranging from 5.0 to 9.0. In scale-up fermentations, the lycopene titer in the engineered strain harboring GPS was increased by 137.3% and ammonia usage decreased by 35.6%. Conclusions The pH self-regulation achieved through the GPS circuits is helpful to construct intelligent microbial cell factories and reduce the production costs, which would be much useful in industrial applications.

Perspectives in growth production trade-off in microbial bioproduction

“…Nitrogen limitation driven dynamic control for itaconate production 41 that was extended to produce itaconate at 1.4 g/L in P. putida from deconstructed lignin. Another example of dynamic control of pH has been reported for lycopene production in E. coli 42 . On-off switches which have been also used by genetic engineers are inversion promoter regions based on recombination events.…”

“…Another example of dynamic control of pH has been reported for the production of lycopene in E. coli . 42 On–off switches, which have been also used by genetic engineers, are inversion promoter regions based on recombination events.…”

Practical genetic control strategies for industrial bioprocesses

“…1 ; Bothfeld et al, 2017 ; Meyer et al, 2018 ), phosphate limitation (Nitta et al, 2021 ; Trung et al, 2019 ), or oxygen supply (Lange et al, 2017 ). Alternatively, pH (Li, C et al, 2020 ) or temperature shift (Wang et al, 2021 ), if consistent with the ideal process and infrastructure, could be employed.…”

Recent advances in engineering of microbial cell factories for intelligent pH regulation and tolerance

“…A genetic pH shooting (GPS) system has been constructed to strengthen the self‐pH‐adjustment ability of E. coli . [ 64 ] The system consisted of a glutaminase gene glsA (catalyzing glutamine to produce ammonia) under the control of the acid‐induced promoter P asr and a lactate dehydrogenase gene ldhA (catalyzing the production of lactic acid) under the control of the base‐induced promoter P atpB from Corynebacterium glutamicum . This system has been shown to significantly improve cell growth and lycopene production while reducing the acid/base usage in the fermenter by stabilizing the environmental pH (Figure 5H).…”