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Continuous fermentation is a key strategy in industrial ethanol production, particularly when efficiency, scalability, and high yields are essential. This approach allows for uninterrupted operation and optimized resource utilization. The primary feedstock, corn starch, undergoes enzymatic hydrolysis facilitated by α-amylase and glucoamylase. These enzymes break down the starch into fermentable sugars such as glucose, which are readily assimilated by fermentative microorganisms.Fermentation...
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A Customizable Approach for the Enzymatic Production and Purification of Diterpenoid Natural Products
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Engineering Rhodosporidium toruloides for limonene production.

Sasa Liu1, Mengyao Zhang1, Yuyao Ren2

  • 1College of Enology, Northwest A&F University, Yangling, Shaanxi, 712100, People's Republic of China.

Biotechnology for Biofuels
|December 23, 2021
PubMed
Summary
This summary is machine-generated.

This study engineered the yeast Rhodosporidium toruloides to produce limonene, a valuable monoterpene. Optimized metabolic pathways and novel enzyme fusions achieved a significant limonene yield of 393.5 mg/L.

Keywords:
LimoneneMetabolic engineeringMicrobial productionMonoterpeneOleaginous yeast

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Area of Science:

  • Microbial biotechnology
  • Synthetic biology
  • Metabolic engineering

Background:

  • Limonene is a versatile monoterpene with broad applications in food, pharmaceuticals, and biofuels.
  • Engineering microbial cell factories is crucial for sustainable production of valuable chemicals.

Purpose of the Study:

  • To engineer the yeast Rhodosporidium toruloides as an efficient cell factory for limonene production.
  • To explore novel metabolic engineering strategies for enhancing limonene biosynthesis in R. toruloides.

Main Methods:

  • Overexpression of key enzymes in the mevalonate pathway, including limonene synthase (LS) and geranyl pyrophosphate synthase (NPPS).
  • Introduction of heterologous genes from Enterococcus faecalis and Methanosarcina mazei to optimize the mevalonate route.
  • Construction of a chimeric NPPS-LS fusion protein and expression in a carotenogenesis-deficient R. toruloides strain.

Main Results:

  • Established a baseline for limonene production in R. toruloides by optimizing the native mevalonate pathway.
  • Achieved a maximum limonene titer of 393.5 mg/L through metabolic engineering and introduction of novel enzymes.
  • Demonstrated the efficacy of the engineered mevalonate pathway and the chimeric NPPS-LS fusion protein.

Conclusions:

  • Successfully engineered R. toruloides for limonene production, marking the first report using this yeast for limonene biosynthesis via the NPP pathway.
  • Validated R. toruloides as a viable microbial platform for the production of limonene and potentially other valuable monoterpenes.
  • Provided valuable insights into the microbial production of monoterpenes through advanced metabolic engineering techniques.