The presence of various furan aldehydes in cellulose hydrolysate affects the fermentation of 1,2,4-butanetriol(BT)in a similar way.In this study,furfural was used as a representative for the modification of BT-produci...The presence of various furan aldehydes in cellulose hydrolysate affects the fermentation of 1,2,4-butanetriol(BT)in a similar way.In this study,furfural was used as a representative for the modification of BT-producing E.coli for tolerance.The engineered Escherichia coli harboring the recombinant BT pathway had decreased the biomass by 58%and the BT titer by 52%in the presence of 0.4 g/L furfural.To improve the tolerance of the strain and the efficiency of BT synthesis in the hydrolysate,seven furfural tolerance genes,ucpA,fucO,groESL,lpcA,pncB,nadD,and nadE were introduced into the BT-producing E.coli.All these genes differentially improved the furfural tolerance performance of the cells.Overexpression of these tolerance genes reduces the accumulation of reactive oxygen species and promotes glycolysis.Oxidoreductase UcpA was the best candidate for improving cell growth.UcpA also increased the activities of Xdh and YqhD and the RNA levels of YjhG and KivD,leading to a 32%increase in BT yield per biomass and the best BT titer of 14.4 g/L in the presence of 0.4 g/L furfural.In the shaker and 5 L fermenter,the BT titer reached 5.2 g/L and 11.2 g/L,respectively,by using corn cob cellulose as substrate.展开更多
Pinene is an active natural monoterpene from plants and has important applications in favorings,fragrances,and pesticides.Especially,pinene dimers are regarded as renewable fuels with high density.However,the microbia...Pinene is an active natural monoterpene from plants and has important applications in favorings,fragrances,and pesticides.Especially,pinene dimers are regarded as renewable fuels with high density.However,the microbial pinene production was limited by the low activity pinene synthase.In this study,the pinene synthase activity was improved by fusion linker optimization and chaperon coexpression.To construct the pinene pathway in Saccharomyces cerevisiae,YPL062W gene was deleted to increase the MVA pathway precursor acetyl-CoA.Truncated 3-hydroxyl-3-methylglutaryl-CoA reductase(tHMG1),isopentenyl-diphosphate isomerase(IDI1),and farnesyl diphosphate synthase mutant(ERG20F96W−N127W)were then integrated to improve the GPP pool.Pinene synthase tPt1 was expressed in the constructed engineered yeast,and the titer of pinene reached 0.166 mg/L.GPP is the direct precursor of pinene,ERG20ww and tPt1 were fused by diferent linkers and orders to improve the accessibility of GPP.Pinene titer reached 9.94 mg/L by fusion these proteins in the order of ERG20ww and tPt1 and with a fexible linker(G)8.After that,several chaperons were coexpressed and the chaperon Sil1p improved the pinene titer to 10.2 mg/L with a yield of 1.63 mg/L·OD600.The results presented here provide novel information on the applications of protein fusion and protein chaperons in microbial pinene production.展开更多
文摘The presence of various furan aldehydes in cellulose hydrolysate affects the fermentation of 1,2,4-butanetriol(BT)in a similar way.In this study,furfural was used as a representative for the modification of BT-producing E.coli for tolerance.The engineered Escherichia coli harboring the recombinant BT pathway had decreased the biomass by 58%and the BT titer by 52%in the presence of 0.4 g/L furfural.To improve the tolerance of the strain and the efficiency of BT synthesis in the hydrolysate,seven furfural tolerance genes,ucpA,fucO,groESL,lpcA,pncB,nadD,and nadE were introduced into the BT-producing E.coli.All these genes differentially improved the furfural tolerance performance of the cells.Overexpression of these tolerance genes reduces the accumulation of reactive oxygen species and promotes glycolysis.Oxidoreductase UcpA was the best candidate for improving cell growth.UcpA also increased the activities of Xdh and YqhD and the RNA levels of YjhG and KivD,leading to a 32%increase in BT yield per biomass and the best BT titer of 14.4 g/L in the presence of 0.4 g/L furfural.In the shaker and 5 L fermenter,the BT titer reached 5.2 g/L and 11.2 g/L,respectively,by using corn cob cellulose as substrate.
基金This work was supported by the National Natural Science Foundation of China(No.31970033).
文摘Pinene is an active natural monoterpene from plants and has important applications in favorings,fragrances,and pesticides.Especially,pinene dimers are regarded as renewable fuels with high density.However,the microbial pinene production was limited by the low activity pinene synthase.In this study,the pinene synthase activity was improved by fusion linker optimization and chaperon coexpression.To construct the pinene pathway in Saccharomyces cerevisiae,YPL062W gene was deleted to increase the MVA pathway precursor acetyl-CoA.Truncated 3-hydroxyl-3-methylglutaryl-CoA reductase(tHMG1),isopentenyl-diphosphate isomerase(IDI1),and farnesyl diphosphate synthase mutant(ERG20F96W−N127W)were then integrated to improve the GPP pool.Pinene synthase tPt1 was expressed in the constructed engineered yeast,and the titer of pinene reached 0.166 mg/L.GPP is the direct precursor of pinene,ERG20ww and tPt1 were fused by diferent linkers and orders to improve the accessibility of GPP.Pinene titer reached 9.94 mg/L by fusion these proteins in the order of ERG20ww and tPt1 and with a fexible linker(G)8.After that,several chaperons were coexpressed and the chaperon Sil1p improved the pinene titer to 10.2 mg/L with a yield of 1.63 mg/L·OD600.The results presented here provide novel information on the applications of protein fusion and protein chaperons in microbial pinene production.
