Production of natural products through metabolic engineering of Saccharomyces cerevisiae

Krivoruchko, Anastasia; Nielsen, Jens

doi:10.1016/j.copbio.2014.12.004

Cited by 180 publications

(121 citation statements)

References 58 publications

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“…However, in order to achieve this goal, greater understanding of the biology underlying chaotrope stress is required. As discussed by Krivoruchko and Nielsen [141], inadequate knowledge of biosynthetic pathways has hindered attempts to metabolically engineer S. cerevisiae to obtain novel products. In relation to optimization of biofuel fermentations (and, indeed, for synthesis of other products), it may also be that adequate progress can only be made by understanding and manipulating biophysical constraints on system function.…”

Section: Discussionmentioning

confidence: 99%

Chaotropicity: a key factor in product tolerance of biofuel-producing microorganisms

Cray

Stevenson

Ball

et al. 2015

Current Opinion in Biotechnology

169

175

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Section: Discussionmentioning

confidence: 99%

Chaotropicity: a key factor in product tolerance of biofuel-producing microorganisms

Cray

Stevenson

Ball

et al. 2015

Current Opinion in Biotechnology

169

175

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“…Optimizing nepetalactol production in yeast is therefore a crucial requisite for realizing microbial production of a wide array of high value compounds including iridoids, strictosidine, as well as all downstream MIA natural products. Accordingly, the recent surge in metabolic engineering technologies in yeast has invigorated natural product pathway engineering efforts (Billingsley et al, 2016; Krivoruchko and Nielsen, 2015). …”

Section: Introductionmentioning

confidence: 99%

Engineering the biocatalytic selectivity of iridoid production in Saccharomyces cerevisiae

Billingsley

DeNicola

Barber

et al. 2017

Metabolic Engineering

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Monoterpene indole alkaloids (MIAs) represent a structurally diverse, medicinally essential class of plant derived natural products. The universal MIA building block strictosidine was recently produced in the yeast Saccharomyces cerevisiae, setting the stage for optimization of microbial production. However, the irreversible reduction of pathway intermediates by yeast enzymes results in a non-recoverable loss of carbon, which has a strong negative impact on metabolic flux. In this study, we identified and engineered the determinants of biocatalytic selectivity which control flux towards the iridoid scaffold from which all MIAs are derived. Development of a bioconversion based production platform enabled analysis of the metabolic flux and interference around two critical steps in generating the iridoid scaffold: oxidation of 8-hydroxygeraniol to the dialdehyde 8-oxogeranial followed by reductive cyclization to form nepetalactol. In vitro reconstitution of previously uncharacterized shunt pathways enabled the identification of two distinct routes to a reduced shunt product including endogenous ‘ene’-reduction and non-productive reduction by iridoid synthase when interfaced with endogenous alcohol dehydrogenases. Deletion of five genes involved in α,β-unsaturated carbonyl metabolism resulted in a 5.2-fold increase in biocatalytic selectivity of the desired iridoid over reduced shunt product. We anticipate that our engineering strategies will play an important role in the development of S. cerevisiae for sustainable production of iridoids and MIAs.

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“…accharomyces cerevisiae has been used as an important cell factory for production of fuels and chemicals (1,2). Although yeast can utilize a wide range of carbon sources, glucose is the most widely preferred carbon source for yeast fermentation (3).…”

mentioning

confidence: 99%

Improvement of Glucose Uptake Rate and Production of Target Chemicals by Overexpressing Hexose Transporters and Transcriptional Activator Gcr1 in Saccharomyces cerevisiae

Kim

Song

Hahn

2015

Appl Environ Microbiol

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Metabolic engineering to increase the glucose uptake rate might be beneficial to improve microbial production of various fuels and chemicals. In this study, we enhanced the glucose uptake rate in Saccharomyces cerevisiae by overexpressing hexose transporters (HXTs). Among the 5 tested HXTs (Hxt1, Hxt2, Hxt3, Hxt4, and Hxt7), overexpression of high-affinity transporter Hxt7 was the most effective in increasing the glucose uptake rate, followed by moderate-affinity transporters Hxt2 and Hxt4. Deletion of STD1 and MTH1, encoding corepressors of HXT genes, exerted differential effects on the glucose uptake rate, depending on the culture conditions. In addition, improved cell growth and glucose uptake rates could be achieved by overexpression of GCR1, which led to increased transcription levels of HXT1 and ribosomal protein genes. All genetic modifications enhancing the glucose uptake rate also increased the ethanol production rate in wild-type S. cerevisiae. Furthermore, the growth-promoting effect of GCR1 overexpression was successfully applied to lactic acid production in an engineered lactic acid-producing strain, resulting in a significant improvement of productivity and titers of lactic acid production under acidic fermentation conditions. Saccharomyces cerevisiae has been used as an important cell factory for production of fuels and chemicals (1, 2). Although yeast can utilize a wide range of carbon sources, glucose is the most widely preferred carbon source for yeast fermentation (3). Glucose uptake in S. cerevisiae is elaborately regulated to adapt cellular metabolism in response to ever-changing environmental conditions (3, 4). However, to achieve high titers and productivity of target chemicals using metabolically engineered yeast cells, it may be beneficial to inactivate, circumvent, or modify some of such natural regulatory mechanisms.The first regulatory step of glucose uptake is the facilitated diffusion of glucose through hexose transporters (HXTs) in the plasma membrane (4, 5). S. cerevisiae contains 18 HXT family members of hexose transporters, Hxt1 to Hxt17 and Gal2, among which Hxt1 to Hxt7 function as major glucose transporters (6). Expression of the HXTs, all of which exhibit different levels of glucose affinity, is differentially regulated depending on extracellular glucose concentrations (7-9). Hxt1 (K m ϭ ϳ100 mM) and Hxt3 (K m ϭ ϳ30 to 60 mM) are classified as low-affinity glucose transporters. HXT1 is highly induced in the presence of high (Ͼ1%) concentrations of glucose, whereas HXT3 is induced by both low and high levels of glucose. When the glucose concentrations are lowered to around 0.1%, HXT2 and HXT4, encoding moderate-affinity glucose transporters (K m ϭ ϳ5 to 10 mM), are expressed (10). HXT6 and HXT7, encoding highly homologous glucose transporters with very high glucose affinity (K m ϭ ϳ1 mM), are expressed just before the depletion of glucose in the medium (11). Expression of HXT5, encoding a moderate-affinity transporter (K m ϭ ϳ10 mM), is not regulated by extracellular glucose ...

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Production of natural products through metabolic engineering of Saccharomyces cerevisiae

Cited by 180 publications

References 58 publications

Chaotropicity: a key factor in product tolerance of biofuel-producing microorganisms

Chaotropicity: a key factor in product tolerance of biofuel-producing microorganisms

Engineering the biocatalytic selectivity of iridoid production in Saccharomyces cerevisiae

Improvement of Glucose Uptake Rate and Production of Target Chemicals by Overexpressing Hexose Transporters and Transcriptional Activator Gcr1 in Saccharomyces cerevisiae

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