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Engineering endogenous ABC transporter with improving ATP supply and membrane flexibility enhances the secretion of β-carotene in Saccharomyces cerevisiae

Authors
  • Bu, Xiao1, 2
  • Lin, Jing-Yuan1, 2
  • Cheng, Jing1, 2
  • Yang, Dong3
  • Duan, Chang-Qing1, 2
  • Koffas, Mattheos4
  • Yan, Guo-Liang1, 2
  • 1 China Agricultural University, 17 East Tsinghua Rd, Beijing, 100083, China , Beijing (China)
  • 2 Ministry of Agriculture and Rural Affairs, Beijing, 100083, China , Beijing (China)
  • 3 China Agricultural University, Beijing, 100083, China , Beijing (China)
  • 4 Rensselaer Polytechnic Institute, Troy, NY, 12180, USA , Troy (United States)
Type
Published Article
Journal
Biotechnology for Biofuels
Publisher
Springer (Biomed Central Ltd.)
Publication Date
Oct 10, 2020
Volume
13
Issue
1
Identifiers
DOI: 10.1186/s13068-020-01809-6
Source
Springer Nature
Keywords
License
Green

Abstract

BackgroundProduct toxicity is one of the bottlenecks for microbial production of biofuels, and transporter-mediated biofuel secretion offers a promising strategy to solve this problem. As a robust microbial host for industrial-scale production of biofuels, Saccharomyces cerevisiae contains a powerful transport system to export a wide range of toxic compounds to sustain survival. The aim of this study is to improve the secretion and production of the hydrophobic product (β-carotene) by harnessing endogenous ABC transporters combined with physiological engineering in S. cerevisiae.ResultsSubstrate inducibility is a prominent characteristic of most endogenous transporters. Through comparative proteomic analysis and transcriptional confirmation, we identified five potential ABC transporters (Pdr5p, Pdr10p, Snq2p, Yor1p, and Yol075cp) for β-carotene efflux. The accumulation of β-carotene also affects cell physiology in various aspects, including energy metabolism, mitochondrial translation, lipid metabolism, ergosterol biosynthetic process, and cell wall synthesis. Here, we adopted an inducible GAL promoter to overexpress candidate transporters and enhanced the secretion and intracellular production of β-carotene, in which Snq2p showed the best performance (a 4.04-fold and a 1.33-fold increase compared with its parental strain YBX-01, respectively). To further promote efflux capacity, two strategies of increasing ATP supply and improving membrane fluidity were following adopted. A 5.80-fold increase of β-carotene secretion and a 1.71-fold increase of the intracellular β-carotene production were consequently achieved in the engineered strain YBX-20 compared with the parental strain YBX-01.ConclusionsOverall, our results showcase that engineering endogenous plasma membrane ABC transporters is a promising approach for hydrophobic product efflux in S. cerevisiae. We also highlight the importance of improving cell physiology to enhance the efficiency of ABC transporters, especially energy status and cell membrane properties.

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