caa a22 4500 605499977 CHVBK 20210128100553.0 cr unu---uuuuu 210128e20150501xx s 000 0 eng 10.1007/s00253-015-6463-y doi (NATIONALLICENCE)springer-10.1007/s00253-015-6463-y Experimental design-aided systematic pathway optimization of glucose uptake and deoxyxylulose phosphate pathway for improved amorphadiene production [Elektronische Daten] [Congqiang Zhang, Ruiyang Zou, Xixian Chen, Gregory Stephanopoulos, Heng-Phon Too] Artemisinin is a potent antimalarial drug; however, it suffers from unstable and insufficient supply from plant source. Here, we established a novel multivariate-modular approach based on experimental design for systematic pathway optimization that succeeded in improving the production of amorphadiene (AD), the precursor of artemisinin, in Escherichia coli. It was initially found that the AD production was limited by the imbalance of glyceraldehyde 3-phosphate (GAP) and pyruvate (PYR), the two precursors of the 1-deoxy-d-xylulose-5-phosphate (DXP) pathway. Furthermore, it was identified that GAP and PYR could be balanced by replacing the phosphoenolpyruvate (PEP)-dependent phosphotransferase system (PTS) with the ATP-dependent galactose permease and glucose kinase system (GGS) and this resulted in fivefold increase in AD titer (11 to 60mg/L). Subsequently, the experimental design-aided systematic pathway optimization (EDASPO) method was applied to systematically optimize the transcriptional expressions of eight critical genes in the glucose uptake and the DXP and AD synthesis pathways. These genes were classified into four modules and simultaneously controlled by T7 promoter or its variants. A regression model was generated using the four-module experimental data and predicted the optimal expression ratios among these modules, resulting in another threefold increase in AD titer (60 to 201mg/L). This EDASPO method may be useful for the optimization of other pathways and products beyond the scope of this study. Springer-Verlag Berlin Heidelberg, 2015 Amorphadiene nationallicence Experimental design-aided systematic pathway optimization nationallicence Multivariate-modular approach nationallicence Deoxyxylulose phosphate pathway nationallicence The phosphotransferase system nationallicence Zhang Congqiang Chemical and Pharmaceutical Engineering, Singapore-MIT Alliance, 4 Engineering Drive 3, Singapore, Singapore aut Zou Ruiyang Chemical and Pharmaceutical Engineering, Singapore-MIT Alliance, 4 Engineering Drive 3, Singapore, Singapore aut Chen Xixian Chemical and Pharmaceutical Engineering, Singapore-MIT Alliance, 4 Engineering Drive 3, Singapore, Singapore aut Stephanopoulos Gregory Chemical and Pharmaceutical Engineering, Singapore-MIT Alliance, 4 Engineering Drive 3, Singapore, Singapore aut Too Heng-Phon Chemical and Pharmaceutical Engineering, Singapore-MIT Alliance, 4 Engineering Drive 3, Singapore, Singapore aut Applied Microbiology and Biotechnology Springer Berlin Heidelberg 99/9(2015-05-01), 3825-3837 0175-7598 99:9<3825 2015 99 253 https://doi.org/10.1007/s00253-015-6463-y text/html Onlinezugriff via DOI BK010053 XK010053 XK010000 Metadata rights reserved Springer special CC-BY-NC licence nationallicence 1 research-article jats NATIONALLICENCE NATIONALLICENCE NL-springer NATIONALLICENCE 856 40 https://doi.org/10.1007/s00253-015-6463-y text/html Onlinezugriff via DOI NATIONALLICENCE 700 1- Zhang Congqiang Chemical and Pharmaceutical Engineering, Singapore-MIT Alliance, 4 Engineering Drive 3, Singapore, Singapore aut NATIONALLICENCE 700 1- Zou Ruiyang Chemical and Pharmaceutical Engineering, Singapore-MIT Alliance, 4 Engineering Drive 3, Singapore, Singapore aut NATIONALLICENCE 700 1- Chen Xixian Chemical and Pharmaceutical Engineering, Singapore-MIT Alliance, 4 Engineering Drive 3, Singapore, Singapore aut NATIONALLICENCE 700 1- Stephanopoulos Gregory Chemical and Pharmaceutical Engineering, Singapore-MIT Alliance, 4 Engineering Drive 3, Singapore, Singapore aut NATIONALLICENCE 700 1- Too Heng-Phon Chemical and Pharmaceutical Engineering, Singapore-MIT Alliance, 4 Engineering Drive 3, Singapore, Singapore aut NATIONALLICENCE 773 0- Applied Microbiology and Biotechnology Springer Berlin Heidelberg 99/9(2015-05-01), 3825-3837 0175-7598 99:9<3825 2015 99 253