Engineering dynamic pathway regulation using stress-response promoters

• Published in: Nature biotechnology Vol. 31; no. 11; pp. 1039 - 1046
• Main Authors: Dahl, Robert H, Zhang, Fuzhong, Alonso-Gutierrez, Jorge, Baidoo, Edward, Batth, Tanveer S, Redding-Johanson, Alyssa M, Petzold, Christopher J, Mukhopadhyay, Aindrila, Lee, Taek Soon, Adams, Paul D, Keasling, Jay D
• Format: Journal Article
• Language: English
• Published: London Nature Publishing Group UK 01.11.2013; Nature Publishing Group
• Subjects: 631/553/2393; 631/553/318; 631/61/185; 631/61/318; Accumulation; Agriculture; Backup software; Bioinformatics; Biomedical Engineering/Biotechnology; Biomedicine; Biosynthesis; Biotechnology; Cluster Analysis; E coli; Enzymes; Escherichia coli; Escherichia coli - genetics; Escherichia coli - physiology; Escherichia coli Proteins - analysis; Escherichia coli Proteins - genetics; Escherichia coli Proteins - metabolism; Gene Expression Profiling - methods; Genes; Genetic regulation; Genomes; Genomics; Instrument industry (Equipment); Keasling, Jay; Life Sciences; Metabolic disorders; Metabolic Engineering - methods; Metabolic Networks and Pathways - genetics; Metabolites; Novels; Phosphates; Polyisoprenyl Phosphates; Promoters (Genetics); Sensors; Sesquiterpenes - metabolism; Stress response; Stress, Physiological - genetics; Systems Biology - methods; United States
• ISSN: 1087-0156; 1546-1696
• DOI: 10.1038/nbt.2689

The problem of toxic intermediates in engineered metabolic pathways is mitigated by dynamic gene-expression regulation using stress-responsive promoters. Heterologous pathways used in metabolic engineering may produce intermediates toxic to the cell. Dynamic control of pathway enzymes could prevent the accumulation of these metabolites, but such a strategy requires sensors, which are largely unknown, that can detect and respond to the metabolite. Here we applied whole-genome transcript arrays to identify promoters that respond to the accumulation of toxic intermediates, and then used these promoters to control accumulation of the intermediate and improve the final titers of a desired product. We apply this approach to regulate farnesyl pyrophosphate (FPP) production in the isoprenoid biosynthetic pathway in Escherichia coli . This strategy improved production of amorphadiene, the final product, by twofold over that from inducible or constitutive promoters, eliminated the need for expensive inducers, reduced acetate accumulation and improved growth. We extended this approach to another toxic intermediate to demonstrate the broad utility of identifying novel sensor-regulator systems for dynamic regulation.