Engineering of the terpenoid pathway in Saccharomyces cerevisiae co-overproduces squalene and the non-terpenoid compound oleic acid

Rasool, Aamir; Zhang, Genlin; Li, Zhe; Li, Chun

doi:10.1016/j.ces.2016.06.004

Cited by 15 publications

(8 citation statements)

References 42 publications

(39 reference statements)

Supporting

Mentioning

Contrasting

Order By: Relevance

“…(A) Squalene biosynthesis via MVA pathway in yeast, fungi, and algae. The engineering strategies for enhanced squalene production are as follows: overexpression of HMGR (Polakowski et al, 1998; Tokuhiro et al, 2009; Mantzouridou and Tsimidou, 2010; Dai et al, 2012, 2014; Zhuang and Chappell, 2015; Rasool et al, 2016a,b; Kwak et al, 2017; Paramasivan and Mutturi, 2017; Han et al, 2018; Huang et al, 2018; Wei et al, 2018) and SQS (Dai et al, 2014; Zhuang and Chappell, 2015; Rasool et al, 2016a,b), downregulation of SQE (Garaiová et al, 2014; Hull et al, 2014; Zhuang and Chappell, 2015; Rasool et al, 2016a,b; Han et al, 2018) in yeast; downregulation of SQE in algae (Kajikawa et al, 2015). (B) Squalene biosynthesis via MEP pathway in bacteria.…”

Section: Squalene Biosynthetic Pathway In Microorganismsmentioning

confidence: 99%

“…Strength of the promoter also plays a crucial role for balancing, tuning and optimizing the expression of a gene toward enhancing the metabolite concentrations. In this perspective, Rasool et al (2016b) overexpressed the genes present in the squalene biosynthetic pathway with a newly characterized and optimized library of 13 new constitutive promoters in S. cerevisiae INVSc1. The resultant engineered strain FOH-0 produced up to 100 mg/L squalene that was 29.41-fold higher than the control strain.…”

Section: Engineering Of Microorganisms For Squalene Productionmentioning

confidence: 99%

See 1 more Smart Citation

Engineering Strategies in Microorganisms for the Enhanced Production of Squalene: Advances, Challenges and Opportunities

Gohil

Bhattacharjee

Khambhati

et al. 2019

Front. Bioeng. Biotechnol.

View full text Add to dashboard Cite

The triterpene squalene is a natural compound that has demonstrated an extraordinary diversity of uses in pharmaceutical, nutraceutical, and personal care industries. Emboldened by this range of uses, novel applications that can gain profit from the benefits of squalene as an additive or supplement are expanding, resulting in its increasing demand. Ever since its discovery, the primary source has been the deep-sea shark liver, although recent declines in their populations and justified animal conservation and protection regulations have encouraged researchers to identify a novel route for squalene biosynthesis. This renewed scientific interest has profited from immense developments in synthetic biology, which now allows fine-tuning of a wider range of plants, fungi, and microorganisms for improved squalene production. There are numerous naturally squalene producing species and strains; although they generally do not make commercially viable yields as primary shark liver sources can deliver. The recent advances made toward improving squalene output from natural and engineered species have inspired this review. Accordingly, it will cover in-depth knowledge offered by the studies of the natural sources, and various engineering-based strategies that have been used to drive the improvements in the pathways toward large-scale production. The wide uses of squalene are also discussed, including the notable developments in anti-cancer applications and in augmenting influenza vaccines for greater efficacy.

show abstract

Section: Squalene Biosynthetic Pathway In Microorganismsmentioning

confidence: 99%

Section: Engineering Of Microorganisms For Squalene Productionmentioning

confidence: 99%

Engineering Strategies in Microorganisms for the Enhanced Production of Squalene: Advances, Challenges and Opportunities

Gohil

Bhattacharjee

Khambhati

et al. 2019

Front. Bioeng. Biotechnol.

View full text Add to dashboard Cite

show abstract

“…The previous report has shown that strength of strong constitutive promoter (GPDp) and strong inducible promoters (GALp) can be further enhanced by adding the enhancer element sequences upstream of core promoter acting as synthetic transcriptional amplifiers [22]. In this study, the strength of previously characterized constitutive promoters is tuned by adding the TFBS from strong promoters (Figure 2a) [14]. Transcription factor binding sequences from TEF1p (−300 to −579 bp) and HHF2p (−300 to −669 bp) promoters were fused upstream of HHF2p, IRA1p, RHO1p, PET9p, CMD1p, ATP16p, USA3p, RER2p, COQ1p, RIM1p, GRS1p, MAK5p and BRN1p (Figure 2b,c, Table S6) and resulting promoters have been listed in the Table S6.…”

Section: Tuning the Strength Of Yeast Promotersmentioning

confidence: 88%

“…This makes it an ideal candidate for industrial-scale production of squalene. Earlier, our engineered S. cerevisiae produced~304.16 mg/L squalene in the shake flask using terbinafine, an inhibitor of squalene epoxidase [14] and synergistically downregulated the expression of ethanol production pathway [15]. This indicates that yeast can self-redirect the metabolic flux from a non-essential pathway to an engineered pathway to alleviate the metabolic burden on pathways critical for its growth [15].…”

Section: Introductionmentioning

confidence: 86%

“…Promoter engineering is a useful strategy adopted to optimize the expression of genes of the engineered pathway for the overproduction of high-value compounds [22]. Therefore, the strength of our earlier characterized 13 yeast constitutive promoters was tuned using transcription factor binding sites (TFBS) from strong promoters HHF2p (−300 to −669 bp) and TEF1p (−300 to −579 bp) and subsequently employed to overexpress the squalene and ethanol production pathways in S. cerevisiae [14].…”

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

Self-Redirection of Metabolic Flux toward Squalene and Ethanol Pathways by Engineered Yeast

et al. 2020

Self Cite

View full text Add to dashboard Cite

We have previously reported that squalene overproducing yeast self-downregulate the expression of the ethanol pathway (non-essential pathway) to divert the metabolic flux to the squalene pathway. In this study, the effect of co-production of squalene and ethanol on other non-essential pathways (fusel alcohol pathway, FA) of Saccharomyces cerevisiae was evaluated. However, before that, 13 constitutive promoters, like IRA1p, PET9p, RHO1p, CMD1p, ATP16p, USA3p, RER2p, COQ1p, RIM1p, GRS1p, MAK5p, and BRN1p, were engineered using transcription factor bindings sites from strong promoters HHF2p (−300 to −669 bp) and TEF1p (−300 to −579 bp), and employed to co-overexpress squalene and ethanol pathways in S. cerevisiae. The FSE strain overexpressing the key genes of the squalene pathway accumulated 56.20 mg/L squalene, a 16.43-fold higher than wild type strain (WS). The biogenesis of lipid droplets was stimulated by overexpressing DGA1 and produced 106 mg/L squalene in the FSE strain. AFT1p and CTR1p repressible promoters were also characterized and employed to downregulate the expression of ERG1, which also enhanced the production of squalene in FSE strain up to 42.85- (148.67 mg/L) and 73.49-fold (255.11 mg/L) respectively. The FSE strain was further engineered by overexpressing the key genes of the ethanol pathway and produced 40.2 mg/mL ethanol in the FSE1 strain, 3.23-fold higher than the WS strain. The FSE1 strain also self-downregulated the expression of the FA pathway up to 73.9%, perhaps by downregulating the expression of GCN4 by 2.24-fold. We demonstrate the successful tuning of the strength of yeast promoters and highest coproduction of squalene and ethanol in yeast, and present GCN4 as a novel metabolic regulator that can be manipulated to divert the metabolic flux from the non-essential pathway to engineered pathways.

show abstract

Improving squalene production by blocking the competitive branched pathways and expressing rate‐limiting enzymes in Rhodopseudomonas palustris

Wang

Fan

et al. 2021

Biotech and App Biochem

View full text Add to dashboard Cite

Squalene is a medically valuable bioactive compound that can be used as a raw material for fuels. Microbial fermentation is the preferred method for the squalene production. In this study, we employed several metabolic engineering strategies to increase squalene yield in Rhodopseudomonas palustris. A 57% increase in squalene titer was achieved by blocking the carotenoid pathway, thus directing more FPP into the squalene biosynthetic pathway. In order to cut down the conversion of squalene to haponoids, a recombinant strain R. palustris [Δshc, ΔcrtB] in which both carotenoid and haponoid pathways were blocked was then constructed, resulting in a 50-fold increase in squalene titer. Based on the expression of rate-limiting enzymes involved in the squalene pathway, the final squalene content reached 23.3 mg/g DCW, which was 178-times higher than that of the wild-type strain. In this study, several methods effective in improving squalene yield have been described and the potential of R. palustris for producing squalene has been demonstrated.

show abstract

Engineering of the terpenoid pathway in Saccharomyces cerevisiae co-overproduces squalene and the non-terpenoid compound oleic acid

Cited by 15 publications

References 42 publications

Engineering Strategies in Microorganisms for the Enhanced Production of Squalene: Advances, Challenges and Opportunities

Engineering Strategies in Microorganisms for the Enhanced Production of Squalene: Advances, Challenges and Opportunities

Self-Redirection of Metabolic Flux toward Squalene and Ethanol Pathways by Engineered Yeast

Improving squalene production by blocking the competitive branched pathways and expressing rate‐limiting enzymes in Rhodopseudomonas palustris

Contact Info

Product

Resources

About