Reconstruction and Evaluation of the Synthetic Bacterial MEP Pathway in Saccharomyces cerevisiae

Partow, Siavash; Siewers, Verena; Daviet, Laurent; Schalk, Michel; Nielsen, Jens

doi:10.1371/journal.pone.0052498

Cited by 59 publications

(39 citation statements)

References 73 publications

(112 reference statements)

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“…Given the variety and essentiality of isoprenoid products in P. falciparum, it is likely that isoprenoid synthesis by the MEP pathway is regulated to control the product availability of IPP and DMAPP. Two ATPs and 3 NADPHs are consumed for the production of each IPP molecule from glucose (142). The cell likely regulates this pathway to optimize energy consumption.…”

Section: Regulation Of Isoprenoid Synthesismentioning

confidence: 99%

Isoprenoid Biosynthesis in Plasmodium falciparum

2014

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bMalaria kills nearly 1 million people each year, and the protozoan parasite Plasmodium falciparum has become increasingly resistant to current therapies. Isoprenoid synthesis via the methylerythritol phosphate (MEP) pathway represents an attractive target for the development of new antimalarials. The phosphonic acid antibiotic fosmidomycin is a specific inhibitor of isoprenoid synthesis and has been a helpful tool to outline the essential functions of isoprenoid biosynthesis in P. falciparum. Isoprenoids are a large, diverse class of hydrocarbons that function in a variety of essential cellular processes in eukaryotes. In P. falciparum, isoprenoids are used for tRNA isopentenylation and protein prenylation, as well as the synthesis of vitamin E, carotenoids, ubiquinone, and dolichols. Recently, isoprenoid synthesis in P. falciparum has been shown to be regulated by a sugar phosphatase. We outline what is known about isoprenoid function and the regulation of isoprenoid synthesis in P. falciparum, in order to identify valuable directions for future research.

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Section: Regulation Of Isoprenoid Synthesismentioning

confidence: 99%

Isoprenoid Biosynthesis in Plasmodium falciparum

2014

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“…Screening for DXP pathway functionality via resistance to the HMGR inhibitor lovastatin was deemed to be unreliable and subject to false positives, likely due to acquired resistance, as reported previously (Partow et al, 2012). To provide a definitive metric for DXP functionality, deletion of one of the first three steps in the mevalonate pathway in S. cerevisiae was considered the best approach since it enables screening for mevalonate-independent growth.…”

Section: Resultsmentioning

confidence: 93%

“…In the first report, seven genes from E. coli encoding the core DXP pathway enzymes were expressed in S. cerevisiae and the pathway was deemed to be functional based on growth of this strain in the presence of lovastatin, an inhibitor of the mevalonate pathway enzyme 3-hydroxy-3-methyl-glutaryl-CoA (HMG-CoA) reductase (Maury et al, 2008). However, a follow-up study by the same group demonstrated that insensitivity to lovastatin was insufficient evidence for DXP pathway functionality since it was not possible to complement a knockout of the mevalonate pathway gene ERG13 , encoding HMG-CoA synthase (Partow et al, 2012). Soon afterwards, an independent study confirmed that expression of the E. coli DXP pathway genes was insufficient to complement an ERG13 knockout in S. cerevisiae (Carlsen et al, 2013).…”

Section: Introductionmentioning

confidence: 99%

“…Despite co-expression of erpA and the redox partners, fldA and fpr , Partow et al found the E. coli IspG to be non-functional in S. cerevisiae (Partow et al, 2012). Faced with a similar result, having expressed these three genes together with the entire ISC machinery from E. coli , Carlsen et al decided to investigate whether an alternative bacterial [4Fe-4S] enzyme could be functionally expressed in the yeast cytosol (Carlsen et al, 2013).…”

Section: Introductionmentioning

confidence: 99%

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Engineering a functional 1-deoxy-D-xylulose 5-phosphate (DXP) pathway in Saccharomyces cerevisiae

Kirby

Dietzel

Wichmann

et al. 2016

Metabolic Engineering

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Isoprenoids are used in many commercial applications and much work has gone into engineering microbial hosts for their production. Isoprenoids are produced either from acetyl-CoA via the mevalonate pathway or from pyruvate and glyceraldehyde 3-phosphate via the 1-deoxy-D-xylulose 5-phosphate (DXP) pathway. Saccharomyces cerevisiae exclusively utilizes the mevalonate pathway to synthesize native isoprenoids and in fact the alternative DXP pathway has never been found or successfully reconstructed in the eukaryotic cytosol. There are, however, several advantages to isoprenoid synthesis via the DXP pathway, such as a higher theoretical yield, and it has long been a goal to transplant the pathway into yeast. In this work, we investigate and address barriers to DXP pathway functionality in S. cerevisiae using a combination of synthetic biology, biochemistry and metabolomics. We report, for the first time, functional expression of the DXP pathway in S. cerevisiae. Under low aeration conditions, an engineered strain relying solely on the DXP pathway for isoprenoid biosynthesis achieved an endpoint biomass 80% of that of the same strain using the mevalonate pathway.

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“…Overexpression of key regulatory enzymes of the native DXP pathway has been also utilized to enhance IPP and DMAPP production in prokaryotes (13,(34)(35)(36)(37). Similarly, in the eukaryotic system, efforts have been made to enhance isoprenoid production via (i) bypassing the endogenous regulation of the MVA pathway in Saccharomyces cerevisiae (38,39) or (ii) modulating the expression of endogenous/homologous enzymes for higher production of isoprenoid-based biofuels in yeast (11,(40)(41)(42). The metabolic engineering efforts for either endogenous or heterologous gene expression have increased the production of a vast array of isoprenoid-based biofuels or fuel precursors.…”

Section: Introductionmentioning

confidence: 99%

Isoprenoid-Based Biofuels: Homologous Expression and Heterologous Expression in Prokaryotes

Phulara

Chaturvedi

Gupta

2016

Appl Environ Microbiol

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bEnthusiasm for mining advanced biofuels from microbial hosts has increased remarkably in recent years. Isoprenoids are one of the highly diverse groups of secondary metabolites and are foreseen as an alternative to petroleum-based fuels. Most of the prokaryotes synthesize their isoprenoid backbone via the deoxyxylulose-5-phosphate pathway from glyceraldehyde-3-phosphate and pyruvate, whereas eukaryotes synthesize isoprenoids via the mevalonate pathway from acetyl coenzyme A (acetyl-CoA). Microorganisms do not accumulate isoprenoids in large quantities naturally, which restricts their application for fuel purposes. Various metabolic engineering efforts have been utilized to overcome the limitations associated with their natural and nonnatural production. The introduction of heterologous pathways/genes and overexpression of endogenous/homologous genes have shown a remarkable increase in isoprenoid yield and substrate utilization in microbial hosts. Such modifications in the hosts' genomes have enabled researchers to develop commercially competent microbial strains for isoprenoid-based biofuel production utilizing a vast array of substrates. The present minireview briefly discusses the recent advancement in metabolic engineering efforts in prokaryotic hosts for the production of isoprenoid-based biofuels, with an emphasis on endogenous, homologous, and heterologous expression strategies.

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Reconstruction and Evaluation of the Synthetic Bacterial MEP Pathway in Saccharomyces cerevisiae

Cited by 59 publications

References 73 publications

Isoprenoid Biosynthesis in Plasmodium falciparum

Isoprenoid Biosynthesis in Plasmodium falciparum

Engineering a functional 1-deoxy-D-xylulose 5-phosphate (DXP) pathway in Saccharomyces cerevisiae

Isoprenoid-Based Biofuels: Homologous Expression and Heterologous Expression in Prokaryotes

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