ArticleNature protocols2014
Production and quantification of sesquiterpenes in Saccharomyces cerevisiae, including extraction, detection and quantification of terpene products and key related metabolites.
Article in Nature protocols, 2014. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 41 papers.
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Who cites it
41 citing papers in PubMed, 83 citations in OpenAlex.
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- Novel cell factory for the production of 24-epi-ergosterol, an un-natural semi-synthetic precursor for the production of brassinolide in Yarrowia lipolytica.World journal of microbiology & biotechnology · 2025Article
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- Fine-Tuning the Function of Farnesene Synthases for Selective Synthesis of Farnesene Stereoisomers.Journal of agricultural and food chemistry · 2024Article
- High-level biosynthesis of enantiopure germacrene D in yeast.Applied microbiology and biotechnology · 2024Article
- Research Progress on Sesquiterpene Compounds from Artabotrys Plants of Annonaceae.Molecules (Basel, Switzerland) · 2024Review
- Product Profiles of Promiscuous Enzymes Can be Altered by Controlling In Vivo Spatial Organization.Advanced science (Weinheim, Baden-Wurttemberg, Germany) · 2023Article
- Molecular and Functional Analyses of Characterized Sesquiterpene Synthases in Mushroom-Forming Fungi.Journal of fungi (Basel, Switzerland) · 2023Article
- Heterologous Production of Artemisinin inBio-protocol · 2023Article
- Quantifying Isoprenoids in the Ergosterol Biosynthesis by Gas Chromatography-Mass Spectrometry.Journal of fungi (Basel, Switzerland) · 2023Article
- Design of Four Small-Molecule-Inducible Systems in the Yeast Chromosome, Applied to Optimize Terpene Biosynthesis.ACS synthetic biology · 2023Article
- Manipulation of sterol homeostasis for the production of 24-epi-ergosterol in industrial yeast.Nature communications · 2023Article
- Functional mining of novel terpene synthases from metagenomes.Biotechnology for biofuels and bioproducts · 2022Article
- Adaptive laboratory evolution of β-caryophyllene producing Saccharomyces cerevisiae.Microbial cell factories · 2021Article
- Auxin-mediated protein depletion for metabolic engineering in terpene-producing yeast.Nature communications · 2021Article
- Heterologous Production of β-Caryophyllene and Evaluation of Its Activity against Plant Pathogenic Fungi.Microorganisms · 2021Article
- Combined Biosynthetic Pathway Engineering and Storage Pool Expansion for High-Level Production of Ergosterol in IndustrialFrontiers in bioengineering and biotechnology · 2021Article
- Combinatorial Metabolic Engineering inBioengineering (Basel, Switzerland) · 2020Article
Corrections and comments
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Authors and funding
4 authors at 2 institutions in 1 country.
Funding
No grant is acknowledged in the PubMed record.
Abstract
The procedures described here are designed for engineering Saccharomyces cerevisiae to produce sesquiterpenes with an aim to either increase product titers or to simply generate a quantity of product sufficient for identification and/or downstream experimentation. Engineering high-level sesquiterpene production in S. cerevisiae often requires iterations of strain modifications and metabolite analysis. To address the latter, the methods described here were tailored for robust measurement of metabolites that we have found to be fundamental indicators of pathway flux, using only gas chromatography and mass spectrometry (GC-MS) instrumentation. Thus, by focusing on heterologous production of sesquiterpenes via the mevalonate (MEV) pathway in S. cerevisiae, we detail procedures for extraction and detection of the key pathway metabolites MEV, squalene and ergosterol, as well as the farnesyl pyrophosphate (FPP)-derived side products farnesol and nerolidol. Analysis of these compounds is important for quality control, because they are possible indicators of pathway imbalance. As many of the sesquiterpene synthase (STS) genes encountered in nature are of plant origin and often not optimal for expression in yeast, we provide guidelines for designing gene expression cassettes to enable expression in S. cerevisiae. As a case study for these protocols, we have selected the sesquiterpene amorphadiene, native to Artemisia annua and related plants. The analytical steps can be completed within 1-2 working days, and a typical experiment might take 1 week.
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