Production efficiency of the bacterial non-ribosomal peptide indigoidine relies on the respiratory metabolic state in <Emphasis Type="Italic">S. cerevisiae</Emphasis> |
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Authors: | Maren Wehrs Jan-Philip Prahl Jadie Moon Yuchen Li Deepti Tanjore Jay D Keasling Aindrila Mukhopadhyay |
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Institution: | 1.Biological Systems and Engineering Division,Lawrence Berkeley National Laboratory,Berkeley,USA;2.Institut für Genetik,Technische Universit?t Braunschweig,Brunswick,Germany;3.Joint BioEnergy Institute, Lawrence Berkeley National Laboratory,Emeryville,USA;4.Advanced Biofuels and Bioproducts Process Development Unit, Lawrence Berkeley National Laboratory,Emeryville,USA;5.Department of Plant and Microbial Biology,University of California,Berkeley,USA;6.Department of Bioengineering,University of California,Berkeley,USA;7.Department of Chemical and Biomolecular Engineering,University of California,Berkeley,USA;8.The Novo Nordisk Foundation Center for Biosustainability,Technical University of Denmark,Kongens Lyngby,Denmark;9.Synthetic Biochemistry Center, Institute for Synthetic Biology,Shenzhen Institutes for Advanced Technologies,Shenzhen,China;10.Environmental Genomics and Systems Biology Division,Lawrence Berkeley National Laboratory,Berkeley,USA |
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Abstract: | BackgroundBeyond pathway engineering, the metabolic state of the production host is critical in maintaining the efficiency of cellular production. The biotechnologically important yeast Saccharomyces cerevisiae adjusts its energy metabolism based on the availability of oxygen and carbon sources. This transition between respiratory and non-respiratory metabolic state is accompanied by substantial modifications of central carbon metabolism, which impact the efficiency of metabolic pathways and the corresponding final product titers. Non-ribosomal peptide synthetases (NRPS) are an important class of biocatalysts that provide access to a wide array of secondary metabolites. Indigoidine, a blue pigment, is a representative NRP that is valuable by itself as a renewably produced pigment.ResultsSaccharomyces cerevisiae was engineered to express a bacterial NRPS that converts glutamine to indigoidine. We characterize carbon source use and production dynamics, and demonstrate that indigoidine is solely produced during respiratory cell growth. Production of indigoidine is abolished during non-respiratory growth even under aerobic conditions. By promoting respiratory conditions via controlled feeding, we scaled the production to a 2 L bioreactor scale, reaching a maximum titer of 980 mg/L.ConclusionsThis study represents the first use of the Streptomyces lavendulae NRPS (BpsA) in a fungal host and its scale-up. The final product indigoidine is linked to the activity of the TCA cycle and serves as a reporter for the respiratory state of S. cerevisiae. Our approach can be broadly applied to investigate diversion of flux from central carbon metabolism for NRPS and other heterologous pathway engineering, or to follow a population switch between respiratory and non-respiratory modes. |
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