Decompartmentalization of the yeast mitochondrial metabolism to improve chemical production in Issatchenkia orientalis
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ABSTRACT:
INSTRUMENT(S): Orbitrap Exploris 480
ORGANISM(S): Pichia Kudriavzevii (ncbitaxon:4909)
SUBMITTER: 
Huimin Zhao
PROVIDER: MSV000098422 | MassIVE | Wed Jul 02 21:48:00 BST 2025
REPOSITORIES: MassIVE
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Project description:BackgroundWith unique physiochemical environments in subcellular organelles, there has been growing interest in harnessing yeast organelles for bioproduct synthesis. Among these organelles, the yeast mitochondrion has been found to be an attractive compartment for production of terpenoids and branched-chain alcohols, which could be credited to the abundant supply of acetyl-CoA, ATP and cofactors. In this study we explored the mitochondrial potential for production of 3-hydroxypropionate (3-HP) and performed the cofactor engineering and flux control at the acetyl-CoA node to maximize 3-HP synthesis.ResultsMetabolic modeling suggested that the mitochondrion serves as a more suitable compartment for 3-HP synthesis via the malonyl-CoA pathway than the cytosol, due to the opportunity to obtain a higher maximum yield and a lower oxygen consumption. With the malonyl-CoA reductase (MCR) targeted into the mitochondria, the 3-HP production increased to 0.27 g/L compared with 0.09 g/L with MCR expressed in the cytosol. With enhanced expression of dissected MCR enzymes, the titer reached to 4.42 g/L, comparable to the highest titer achieved in the cytosol so far. Then, the mitochondrial NADPH supply was optimized by overexpressing POS5 and IDP1, which resulted in an increase in the 3-HP titer to 5.11 g/L. Furthermore, with induced expression of an ACC1 mutant in the mitochondria, the final 3-HP production reached 6.16 g/L in shake flask fermentations. The constructed strain was then evaluated in fed-batch fermentations, and produced 71.09 g/L 3-HP with a productivity of 0.71 g/L/h and a yield on glucose of 0.23 g/g.ConclusionsIn this study, the yeast mitochondrion is reported as an attractive compartment for 3-HP production. The final 3-HP titer of 71.09 g/L with a productivity of 0.71 g/L/h was achieved in fed-batch fermentations, representing the highest titer reported for Saccharomyces cerevisiae so far, that demonstrated the potential of recruiting the yeast mitochondria for further development of cell factories.
| S-EPMC10082987 | biostudies-literature
Project description:BACKGROUND:Saccharomyces cerevisiae is a suitable host for the industrial production of pyruvate-derived chemicals such as ethanol and 2,3-butanediol (23BD). For the improvement of the productivity of these chemicals, it is essential to suppress the unnecessary pyruvate consumption in S. cerevisiae to redirect the metabolic flux toward the target chemical production. In this study, mitochondrial pyruvate transporter gene (MPC1) or the essential gene for mitophagy (ATG32) was knocked-out to repress the mitochondrial metabolism and improve the production of pyruvate-derived chemical in S. cerevisiae. RESULTS:The growth rates of both aforementioned strains were 1.6-fold higher than that of the control strain. 13C-metabolic flux analysis revealed that both strains presented similar flux distributions and successfully decreased the tricarboxylic acid cycle fluxes by 50% compared to the control strain. Nevertheless, the intracellular metabolite pool sizes were completely different, suggesting distinct metabolic effects of gene knockouts in both strains. This difference was also observed in the test-tube culture for 23BD production. Knockout of ATG32 revealed a 23.6-fold increase in 23BD titer (557.0 ± 20.6 mg/L) compared to the control strain (23.5 ± 12.8 mg/L), whereas the knockout of MPC1 revealed only 14.3-fold increase (336.4 ± 113.5 mg/L). Further investigation using the anaerobic high-density fermentation test revealed that the MPC1 knockout was more effective for ethanol production than the 23BD production. CONCLUSION:These results suggest that the engineering of the mitochondrial transporters and membrane dynamics were effective in controlling the mitochondrial metabolism to improve the productivities of chemicals in yeast cytosol.
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Project description:Contains a collection of wildtype matA Saccharomyces cerevisiae strains for estimating the biological variation.
2012-11-14 | GSE42240 | GEO
Project description:Inorganic-biological hybrid systems have potential to be sustainable, efficient, and versatile chemical synthesis platforms by integrating the light-harvesting properties of semiconductors with the synthetic potential of biological cells. We have developed a modular bioinorganic hybrid platform that consists of highly efficient light-harvesting indium phosphide nanoparticles and genetically engineered Saccharomyces cerevisiae, a workhorse microorganism in biomanufacturing. The yeast harvests photogenerated electrons from the illuminated nanoparticles and uses them for the cytosolic regeneration of redox cofactors. This process enables the decoupling of biosynthesis and cofactor regeneration, facilitating a carbon- and energy-efficient production of the metabolite shikimic acid, a common precursor for several drugs and fine chemicals. Our work provides a platform for the rational design of biohybrids for efficient biomanufacturing processes with higher complexity and functionality.
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Project description:Contains a collection of wildtype matA Saccharomyces cerevisiae strains grown using a Tecan platereader for estimating the biological variation.
2012-11-14 | GSE42241 | GEO
Project description:The many virtues that made the yeast Saccharomyces cerevisiae a dominant model organism for genetics and molecular biology, are now establishing its role in chemical genetics. Its experimental tractability (i.e., rapid doubling time, simple culture conditions) and the availability of powerful tools for drug-target identification, make yeast an ideal organism for high-throughput phenotypic screening. It may be especially applicable for the discovery of chemical probes targeting highly conserved cellular processes, such as metabolism and bioenergetics, because these probes would likely inhibit the same processes in higher eukaryotes (including man). Importantly, changes in normal cellular metabolism are associated with a variety of diseased states (including neurological disorders and cancer), and exploiting these changes for therapeutic purposes has accordingly gained considerable attention. Here, we review progress and challenges associated with forward chemical genetic screening in yeast. We also discuss evidence supporting these screens as a useful strategy for discovery of new chemical probes and new druggable targets related to cellular metabolism.
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Project description:Contains a collection of wildtype Saccharomyces cerevisiae strains for estimating the biological variation. Wildtypes are obtained from the yeast wildtypes - wt pool background set HybSet, by randomly taking 100 wt vs. refpool (pooled wts) and 100 refpool vs. wt hybridizations
2012-11-14 | GSE42215 | GEO
Project description:This review gives an overview of different yeast strains and enzyme classes involved in yeast whole-cell biotransformations. A focus was put on the synthesis of compounds for fine chemical and API (= active pharmaceutical ingredient) production employing single or only few-step enzymatic reactions. Accounting for recent success stories in metabolic engineering, the construction and use of synthetic pathways was also highlighted. Examples from academia and industry and advances in the field of designed yeast strain construction demonstrate the broad significance of yeast whole-cell applications. In addition to Saccharomyces cerevisiae, alternative yeast whole-cell biocatalysts are discussed such as Candida sp., Cryptococcus sp., Geotrichum sp., Issatchenkia sp., Kloeckera sp., Kluyveromyces sp., Pichia sp. (including Hansenula polymorpha = P. angusta), Rhodotorula sp., Rhodosporidium sp., alternative Saccharomyces sp., Schizosaccharomyces pombe, Torulopsis sp., Trichosporon sp., Trigonopsis variabilis, Yarrowia lipolytica and Zygosaccharomyces rouxii.
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Project description:Biosynthetic gene clusters (BGCs) encoding the production of bacteriocins are widespread amongst bacterial isolates and are important genetic determinants of competitive fitness among bacterial lineages within a given habitat. Staphylococci produce a tremendous diversity of compounds and the corresponding gene clusters (Staphylococcal Antibiosis Islands - SAbIs) are frequently associated with mobile genetic elements, suggesting acquisition and loss of biosynthetic capacity. Pharmaceutical biology has shown that compound production in heterologous hosts is often challenged by the lack of optimal precursor supplies. Accordingly, many recipients produce low compound amounts or show reduced growth rates. To assess whether transfer of SAbIs between closely related S. aureus strains has similar effects, we used as model the SAbI encoding the ribosomally synthesized and post-translationally modified peptide (RiPP) Micrococcin P1 (MP1). We found that acquisition of the SAbI by S. aureus RN4220 did allow immediate MP1 production but also imposed a metabolic burden. Adaptive evolution selected for strains with increased TCA-cycle activity, which enhanced metabolic fitness and levels of compound production. Metabolome analysis showed that the adaptive mutation increased the levels of central metabolites including citrate and α-ketoglutarate, and the increased availability of citrate turned out to be essential to overcome SAbI associated growth defects. Our results indicate that acquisition of SAbIs represents a blessing for the recipient strain as long as its genetic and metabolic predispositions allow integration of bacteriocin production into the cellular metabolism. This in turn may be crucial to ensure the competitive fitness of SAbI recipients within natural bacterial communities.
2022-11-21 | GSE208001 | GEO
Project description:We aimed to study how production of p-coumaric acid, a precursor of multiple secondary aromatic metabolites, influences the cellular metabolism of Saccharomyces cerevisiae. We evaluated the growth and p-coumaric acid production in batch and chemostat cultivations and analyzed the transcriptome and intracellular metabolome during steady state in low- and high-producers of p-coumaric acid in two strain backgrounds, S288c or CEN.PK. For analysis of the differential gene expression, we did pairwise comparisons between the optimized and non-optimized strains for p-CA production: CEN.PK strains (ST4288 and ST4408) and the S288c strains (ST4353 and ST4397). Transcriptome analysis showed that the CEN.PK strain was less affected by engineering towards higher p-CA production than the S288c strain, as the number of significantly up-/down-regulated genes was correspondingly 652 and 1927 amongst others, strain S288c had downregulations in gene sets involved in amino acid and protein biosynthesis. This suggests that CEN.PK may be a better platform strain for production of aromatic compounds than the S288c strain.
2017-11-21 | E-MTAB-6208 | biostudies-arrayexpress