Microbial synthesis of carotenoids is a highly desirable alternative to plant extraction and chemical synthesis.In this study,we investigated multidimensional strategies to improve the carotenoid synthesis in the indu...Microbial synthesis of carotenoids is a highly desirable alternative to plant extraction and chemical synthesis.In this study,we investigated multidimensional strategies to improve the carotenoid synthesis in the industrial workhorse of Saccharomyces cerevisiae.First,we rewired the yeast central metabolism by optimizing non-oxidative glycolysis pathway for an improved acetyl-CoA supply.Second,we restricted the consumption of farnesyl pyrophosphate(FPP)by the down-regulation of squalene synthase using the PEST degron.Third,we further explored the human lipid binding/transfer protein saposin B(hSapB)-mediated metabolic sink for an enhanced storage of lipophilic carotenoids.Last,the copper-induced GAL expression system was engineered to function in the yeast-peptone-dextrose medium for an increased biomass accumulation.By combining the abovementioned strategies,the final engineered yeast produced 166.79±10.43 mg/Ⅰβ-carotene in shake flasks,which was nearly 5-fold improvement of the parental carotenoid-producing strain.Together,we envision that multidimensional strategies reported here might be applicable to other hosts for the future industrial development of carotenoid synthesis from renewable feedstocks.展开更多
Extracellular polymeric substances(EPSs)are extracellular macromolecules in bacteria,which function in cell growth and show potential for mechanism study and biosynthesis application.However,the biosynthesis mechanism...Extracellular polymeric substances(EPSs)are extracellular macromolecules in bacteria,which function in cell growth and show potential for mechanism study and biosynthesis application.However,the biosynthesis mechanism of EPS is still not clear.We herein chose Bacillus licheniformis CGMCC 2876 as a target strain to investigate the EPS biosynthesis.epsK,a member of eps cluster,the predicted polysaccharide synthesis cluster,was overexpressed and showed that the overexpression of epsK led to a 26.54%decrease in the production of EPS and resulted in slenderer cell shape.Transcriptome analysis combined with protein-protein interactions analysis and protein modeling revealed that epsK was likely responsible for the synthesis of teichuronic acid,a substitute cell wall component of teichoic acid when the strain was suffering phosphate limitation.Further cell cultivation showed that either phosphate limitation or the overexpression of teichuronic acid synthesis genes,tuaB and tuaE could similarly lead to EPS reduction.The enhanced production of teichuronic acid induced by epsK overexpression triggered the endogenous phosphate starvation,resulting in the decreased EPS synthesis and biomass,and the enhanced bacterial chemotaxis.This study presents an insight into the mechanism of EPS synthesis and offers the potential in controllable synthesis of target products.展开更多
Poly-γ-glutamic acid is an extracellular polymeric substance with various applications owing to its valuable properties of biodegradability,flocculating activity,water solubility,and nontoxicity.However,the ability o...Poly-γ-glutamic acid is an extracellular polymeric substance with various applications owing to its valuable properties of biodegradability,flocculating activity,water solubility,and nontoxicity.However,the ability of natural strains to produce poly-γ-glutamic acid is low.Atmospheric and room temperature plasma was applied in this study to conduct mutation breeding of Bacillus licheniformis CGMCC 2876,and a mutant strain M32 with an 11%increase in poly-γ-glutamic acid was obtained.Genome resequencing analysis identified 7 nonsynonymous mutations of ppsC encoding lipopeptide synthetase associated with poly-γ-glutamic acid metabolic pathways.From molecular docking,more binding sites and higher binding energy were speculated between the mutated plipastatin synthase subunit C and glutamate,which might contribute to the higher poly-γ-glutamic acid production.Moreover,the metabolic mechanism analysis revealed that the upregulated amino acids of M32 provided substrates for glutamate and promoted the conversion between L-and D-glutamate acids.In addition,the glycolytic pathway is enhanced,leading to a better capacity for using glucose.The maximum poly-γ-glutamic acid yield of 14.08 g·L^(–1)was finally reached with 30 g·L^(–1)glutamate.展开更多
基金support from the National Natural Science Foundation of China(32270087)the Natural Science Foundation of Fujian Province of China(2020J05011)+2 种基金Guangdong Basic and Applied Basic Research Foundation(2021A1515110340)Xiamen University(0660X2510200)Daan Gene(20223160A0063),and ZhenSheng Biotech.
文摘Microbial synthesis of carotenoids is a highly desirable alternative to plant extraction and chemical synthesis.In this study,we investigated multidimensional strategies to improve the carotenoid synthesis in the industrial workhorse of Saccharomyces cerevisiae.First,we rewired the yeast central metabolism by optimizing non-oxidative glycolysis pathway for an improved acetyl-CoA supply.Second,we restricted the consumption of farnesyl pyrophosphate(FPP)by the down-regulation of squalene synthase using the PEST degron.Third,we further explored the human lipid binding/transfer protein saposin B(hSapB)-mediated metabolic sink for an enhanced storage of lipophilic carotenoids.Last,the copper-induced GAL expression system was engineered to function in the yeast-peptone-dextrose medium for an increased biomass accumulation.By combining the abovementioned strategies,the final engineered yeast produced 166.79±10.43 mg/Ⅰβ-carotene in shake flasks,which was nearly 5-fold improvement of the parental carotenoid-producing strain.Together,we envision that multidimensional strategies reported here might be applicable to other hosts for the future industrial development of carotenoid synthesis from renewable feedstocks.
基金This work was financially supported by the National Natural Science Foundation of China(31871779 and 32170061).
文摘Extracellular polymeric substances(EPSs)are extracellular macromolecules in bacteria,which function in cell growth and show potential for mechanism study and biosynthesis application.However,the biosynthesis mechanism of EPS is still not clear.We herein chose Bacillus licheniformis CGMCC 2876 as a target strain to investigate the EPS biosynthesis.epsK,a member of eps cluster,the predicted polysaccharide synthesis cluster,was overexpressed and showed that the overexpression of epsK led to a 26.54%decrease in the production of EPS and resulted in slenderer cell shape.Transcriptome analysis combined with protein-protein interactions analysis and protein modeling revealed that epsK was likely responsible for the synthesis of teichuronic acid,a substitute cell wall component of teichoic acid when the strain was suffering phosphate limitation.Further cell cultivation showed that either phosphate limitation or the overexpression of teichuronic acid synthesis genes,tuaB and tuaE could similarly lead to EPS reduction.The enhanced production of teichuronic acid induced by epsK overexpression triggered the endogenous phosphate starvation,resulting in the decreased EPS synthesis and biomass,and the enhanced bacterial chemotaxis.This study presents an insight into the mechanism of EPS synthesis and offers the potential in controllable synthesis of target products.
基金This work was financially supported by the National Natural Science Foundation of China(Grant Nos.32170061 and 31871779).
文摘Poly-γ-glutamic acid is an extracellular polymeric substance with various applications owing to its valuable properties of biodegradability,flocculating activity,water solubility,and nontoxicity.However,the ability of natural strains to produce poly-γ-glutamic acid is low.Atmospheric and room temperature plasma was applied in this study to conduct mutation breeding of Bacillus licheniformis CGMCC 2876,and a mutant strain M32 with an 11%increase in poly-γ-glutamic acid was obtained.Genome resequencing analysis identified 7 nonsynonymous mutations of ppsC encoding lipopeptide synthetase associated with poly-γ-glutamic acid metabolic pathways.From molecular docking,more binding sites and higher binding energy were speculated between the mutated plipastatin synthase subunit C and glutamate,which might contribute to the higher poly-γ-glutamic acid production.Moreover,the metabolic mechanism analysis revealed that the upregulated amino acids of M32 provided substrates for glutamate and promoted the conversion between L-and D-glutamate acids.In addition,the glycolytic pathway is enhanced,leading to a better capacity for using glucose.The maximum poly-γ-glutamic acid yield of 14.08 g·L^(–1)was finally reached with 30 g·L^(–1)glutamate.
