Abstract
Optimization of export mechanisms for valuable extracellular products is important for the development of efficient microbial production processes. Identification of the relevant export mechanism is the prerequisite step for product export optimization. In this work, we identified transporters involved in malate export in an engineered l-malate-producing Escherichia coli strain using cheminformatics-guided genetics tests. Among all short-chain di- or tricarboxylates with known transporters in E. coli, citrate, tartrate, and succinate are most chemically similar to malate as estimated by their molecular signatures. Inactivation of three previously reported transporters for succinate, tartrate, and citrate, DcuA, TtdT, and CitT, respectively, dramatically decreased malate production and fermentative growth, suggesting that these transporters have substrate promiscuity for different short-chain organic acids and constitute the major malate export system in E. coli. Malate export deficiency led to an increase in cell sizes and accumulation of intracellular metabolites related to malate metabolism.
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Acknowledgments
We thank the Ingram laboratory at the University of Florida providing the strain XZ658. We thank Dr. Robert Roberson for technical assistance with the microscopic experiments. We thank members of the Neuer laboratory for assistance with cell density measurements.
Funding
This work was supported by the start-up fund and the LightWorks seed grant from Arizona State University (ASU). Eric Taylor and Aidan Schneider were supported by the SOLUR program of ASU. We appreciate multiple fellowships from Arizona State University and other agencies awarded to Logan Kurgan (the IMSD program and the USE scholarship) and Rodrigo Martinez (WAESO-LSAMP Bridge to Doctorate Fellowship).
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Kurgan, G., Kurgan, L., Schneider, A. et al. Identification of major malate export systems in an engineered malate-producing Escherichia coli aided by substrate similarity search. Appl Microbiol Biotechnol 103, 9001–9011 (2019). https://doi.org/10.1007/s00253-019-10164-y
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DOI: https://doi.org/10.1007/s00253-019-10164-y