An endoplasmic reticulum-engineered yeast platform for overproduction of triterpenoids

Arendt, Philipp; Miettinen, Karel; Pollier, Jacob; Rycke, Riet De; Callewaert, Nico; Goossens, Alain

doi:10.1016/j.ymben.2017.02.007

Cited by 141 publications

(106 citation statements)

References 55 publications

Supporting

Mentioning

102

Contrasting

Order By: Relevance

“…The effects of several previously unexplored gene knockout targets of Saccharomyces cerevisiae were assessed for the heterologous production and to improve the production capabilities of this saponin production platform. Dramatic expansion of the endoplasmic reticulum stimulates the production of recombinant triterpene biosynthetic enzymes through disruption of the phosphatidic acid phosphatase-encoding PAH1 of Saccharomyces cerevisiae through CRISPR/Cas9 which results in the accumulation of triterpenoids and triterpene saponin (Arendt et al 2017). Transgenic yeast strains expressing β-amyrin synthase (bAS), P450 reductase, CyP93E2 and CyP72A61v2 were able to produce soyasapogenol B, and yeast strain over expressing bAS, P450 reductase, CyP716A12 and CyP72A68v2 produce gypsogenic acid.…”

Section: Production Of Steroidal Saponin In Culture Mediamentioning

confidence: 99%

Recent advances in steroidal saponins biosynthesis and in vitro production

Upadhyay

Jeena

Shikha

et al. 2018

Planta

View full text Add to dashboard Cite

Main conclusion Steroidal saponins exhibited numerous pharmacological activities due to the modification of their backbone by different cytochrome P450s (P450) and UDP glycosyltransferases (UGTs). Plant-derived steroidal saponins are not sufficient for utilizing them for commercial purpose so in vitro production of saponin by tissue culture, root culture, embryo culture, etc, is necessary for its large-scale production.Saponin glycosides are the important class of plant secondary metabolites, which consists of either steroidal or terpenoidal backbone. Due to the existence of a wide range of medicinal properties, saponin glycosides are pharmacologically very important. This review is focused on important medicinal properties of steroidal saponin, its occurrence, and biosynthesis. In addition to this, some recently identified plants containing steroidal saponins in different parts were summarized. The high throughput transcriptome sequencing approach elaborates our understanding related to the secondary metabolic pathway and its regulation even in the absence of adequate genomic information of non-model plants. The aim of this review is to encapsulate the information related to applications of steroidal saponin and its biosynthetic enzymes specially P450s and UGTs that are involved at later stage modifications of saponin backbone. Lastly, we discussed the in vitro production of steroidal saponin as the plant-based production of saponin is time-consuming and yield a limited amount of saponins. A large amount of plant material has been used to increase the production of steroidal saponin by employing in vitro culture technique, which has received a lot of attention in past two decades and provides a way to conserve medicinal plants as well as to escape them for being endangered.

show abstract

Section: Production Of Steroidal Saponin In Culture Mediamentioning

confidence: 99%

Recent advances in steroidal saponins biosynthesis and in vitro production

Upadhyay

Jeena

Shikha

et al. 2018

Planta

View full text Add to dashboard Cite

show abstract

“…The consequences of overexpressing other MVA pathway genes were determined by combinatorial library screening for the overexpression of ERG10 (acetoacetyl CoA thiolase; AACT), ERG13 (HMGS), and ERG12 (mevalonate kinase) which enhanced the production of amorpha-4,11-diene (Yuan and Ching 2014). The MVA pathway has also been targeted using the CRISPR/Cas9 system, revealing loci that trigger the accumulation of mevalonate and triterpenes when knocked out (Jakočiūnas et al 2015; Arendt et al 2017). The targets included ROX1 , encoding a transcriptional regulator that inhibits genes involved in the MVA pathway and sterol biosynthesis (Henry et al 2002; Montañés et al 2011; Özaydin et al 2013; Jakočiūnas et al 2015).…”

Section: Introductionmentioning

confidence: 99%

Upregulating the mevalonate pathway and repressing sterol synthesis in Saccharomyces cerevisiae enhances the production of triterpenes

Bröker

Müller

Deenen

et al. 2018

Appl Microbiol Biotechnol

View full text Add to dashboard Cite

Pentacyclic triterpenes are diverse plant secondary metabolites derived from the mevalonate (MVA) pathway. Many of these molecules are potentially valuable, particularly as pharmaceuticals, and research has focused on their production in simpler and more amenable heterologous systems such as the yeast Saccharomyces cerevisiae. We have developed a new heterologous platform for the production of pentacyclic triterpenes in S. cerevisiae based on a combinatorial engineering strategy involving the overexpression of MVA pathway genes, the knockout of negative regulators, and the suppression of a competing pathway. Accordingly, we overexpressed S. cerevisiae ERG13, encoding 3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) synthase, and a truncated and deregulated variant of the rate-limiting enzyme HMG-CoA reductase 1 (tHMGR). In the same engineering step, we deleted the ROX1 gene, encoding a negative regulator of the MVA pathway and sterol biosynthesis, resulting in a push-and-pull strategy to enhance metabolic flux through the system. In a second step, we redirected this enhanced metabolic flux from late sterol biosynthesis to the production of 2,3-oxidosqualene, the direct precursor of pentacyclic triterpenes. In yeast cells transformed with a newly isolated sequence encoding lupeol synthase from the Russian dandelion (Taraxacum koksaghyz), we increased the yield of pentacyclic triterpenes by 127-fold and detected not only high levels of lupeol but also a second valuable pentacyclic triterpene product, β-amyrin.

show abstract

“…Lipid metabolism was proven to be an effective strategy that can relieve the downstream bottleneck of terpene synthesis, including by extending the storage capacity of lipophilic products or improving terpene synthase e ciency, thereby promoting terpene overproduction [9][10][11][12][13]. As such, we manipulated eight structural and regulatory genes related to lipid metabolism in LU-9 ( Fig.…”

Section: Engineering Of Mva and Lipid Metabolism To Improve Lupeol Prmentioning

confidence: 99%

“…For instance, lupeol, a typical triterpenoid, caused severe damage to cell viability even at a relatively low concentration of 60 mg/L [3,8]. To decrease the intractable cytotoxicity, extensive efforts aimed at creating oleaginous subcellular organelles by altering lipid-droplet composition and size potentially improved the terpene partition coe cient in oil droplets and the storage space so that lipophilic terpenes can accumulate in these compartments [9][10][11][12][13][14][15]. For instance, researchers successfully increased lycopene accumulation by creating supersized lipid droplets through manipulation of triacylglycerol (TAG) metabolism in Saccharomyces cerevisiae, which resulted in the highest yield (73.3 mg/g cdw and 2.37 g/L lycopene) reported in S. cerevisiae to date [10].…”

Section: Introductionmentioning

confidence: 99%

High production of triterpenoids in Yarrowia lipolytica through manipulation of lipid components

Zhang

Bai

Peng

et al. 2020

Preprint

View full text Add to dashboard Cite

Background Lupeol exhibits novel physiological and pharmacological activities, such as anticancer and immunity-enhancing activities. However, cytotoxicity remains a challenge for triterpenoid overproduction in microbial cell factories. As lipophilic and relatively small-molecular compounds, triterpenes are generally secreted into the extracellular space. The effect of increasing triterpene efflux on the synthesis capacity remains unknown.Results In this study, we developed a strategy to enhance triterpene efflux through manipulation of lipid components in Y. lipolytica by overexpressing the enzyme Δ9-fatty acid desaturase (OLE1) and disturbing phosphatidic acid phosphatase (PAH1) and diacylglycerol kinase (DGK1). By this strategy combined with two-phase fermentation, the highest lupeol production reported to date was achieved, where the titer in the organic phase reached 381.67 mg/L and the total production was 411.72 mg/L in shake flasks, exhibiting a 33.20-fold improvement over the initial strain. Lipid manipulation led to a two-fold increase in the unsaturated fatty acid (UFA) content, up to 61%~73%, and an exceptionally elongated cell morphology, which might have been caused by enhanced membrane phospholipid biosynthesis flux. Both phenotypes accelerated the export of toxic products to the extracellular space and ultimately stimulated the capacity for triterpenoid synthesis, which was proven by the 5.11-fold higher ratio of extra/intracellular lupeol concentrations, 2.79-fold higher biomass accumulation and 2.56-fold higher lupeol productivity per unit OD in the modified strains. This strategy was also highly efficient for the biosynthesis of other triterpenes and sesquiterpenes, including α-amyrin, β-amyrin, longifolene, longipinene and longicyclene.Conclusions In conclusion, we successfully created a high-yield lupeol-producing strain via lipid manipulation. We demonstrated that the enhancement of lupeol efflux and synthesis capacity was induced by the increased UFA content and elongated cell morphology. Our study provides a novel strategy to promote the biosynthesis of valuable but toxic products in microbial cell factories.

show abstract

An endoplasmic reticulum-engineered yeast platform for overproduction of triterpenoids

Cited by 141 publications

References 55 publications

Recent advances in steroidal saponins biosynthesis and in vitro production

Recent advances in steroidal saponins biosynthesis and in vitro production

Upregulating the mevalonate pathway and repressing sterol synthesis in Saccharomyces cerevisiae enhances the production of triterpenes

High production of triterpenoids in Yarrowia lipolytica through manipulation of lipid components

Contact Info

Product

Resources

About