Proteome reallocation enables the selective de novo biosynthesis of non-linear, branched-chain acetate esters

Seo, Hyung-Min; Giannone, Richard J.; Yang, Yung‐Hun; Trinh, Cong T.

doi:10.1016/j.ymben.2022.05.003

Cited by 8 publications

(21 citation statements)

References 56 publications

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“…Here, we investigated how the CCR-engineered strains modulated Each of these two strains was paired with a glucose-utilizing E. coli specialist partner (i.e., HSEC0302_iLOV) harboring an iLOV expression plasmid that generates GFP signals (Figure 3A). HSEC0302, derived from HSEC0201, lacks L-lactate dehydrogenase (ldhA), 8 which is beneficial for pyruvate-derived biochemical production with reduced lactate formation. For control experiments, the characterization results in monocultures showed that the iLOV (GFP) and mCHERRY (RFP) fluorescence signals did not interfere with each other (Figure 3B), suggesting that monitoring the two fluorescence intensities enables the dissection of two species in the coculture.…”

Section: ■ Results and Discussionmentioning

confidence: 99%

Engineering a Synthetic Escherichia coli Coculture for Compartmentalized de novo Biosynthesis of Isobutyl Butyrate from Mixed Sugars

Seo,

Castro,

Trinh

2023

ACS Synth. Biol.

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Short-chain esters are versatile chemicals that can be used as flavors, fragrances, solvents, and fuels. The de novo ester biosynthesis consists of diverging and converging pathway submodules, which is challenging to engineer to achieve optimal metabolic fluxes and selective product synthesis. Compartmentalizing the pathway submodules into specialist cells that facilitate pathway modularization and labor division is a promising solution. Here, we engineered a synthetic Escherichia coli coculture with the compartmentalized sugar utilization and ester biosynthesis pathways to produce isobutyl butyrate from a mixture of glucose and xylose. To compartmentalize the sugar-utilizing pathway submodules, we engineered a xylose-utilizing E. coli specialist that selectively consumes xylose over glucose and bypasses carbon catabolite repression (CCR) while leveraging the native CCR machinery to activate a glucose-utilizing E. coli specialist. We found that the compartmentalization of sugar catabolism enabled simultaneous co-utilization of glucose and xylose by a coculture of the two E. coli specialists, improving the stability of the coculture population. Next, we modularized the isobutyl butyrate pathway into the isobutanol, butyl-CoA, and ester condensation submodules, where we distributed the isobutanol submodule to the glucose-utilizing specialist and the other submodules to the xylose-utilizing specialist. Upon compartmentalization of the isobutyl butyrate pathway submodules into these sugar-utilizing specialist cells, a robust synthetic coculture was engineered to selectively produce isobutyl butyrate, reduce the biosynthesis of unwanted ester byproducts, and improve the production titer as compared to the monoculture.

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Section: ■ Results and Discussionmentioning

confidence: 99%

Engineering a Synthetic Escherichia coli Coculture for Compartmentalized de novo Biosynthesis of Isobutyl Butyrate from Mixed Sugars

Seo,

Castro,

Trinh

2023

ACS Synth. Biol.

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“…In any case, it is known that, depending on the yeast species involved and the environmental conditions, many VOCs may be obtained from carbohydrate metabolism [ 72 , 73 , 74 ]. This is especially relevant for acetaldehyde, ethanol, ethyl acetate, acetic acid, and acetoin, which can be produced within one-to-three reactions (of a metabolic pathway) from pyruvate [ 75 , 76 , 77 , 78 , 79 , 80 ]. In fact, as an intermediate product of sugar oxidation, pyruvate can work as a wildcard and be used in different metabolic pathways ( Figure 2 ).…”

Section: Yeasts At Work: Nectar Fermentation and Voc Productionmentioning

confidence: 99%

“…In this case, though, it must be first reduced into 2,3-dihydroxyisovalerate, and this is then dehydrated to 2-ketoisovalerate. In turn, 2-ketoisovalerate can be either aminated, producing valine, or decarboxylated into isobutyraldehyde, which is reduced into isobutanol [ 75 , 77 , 80 ]. Finally, pyruvate can also be oxidized and decarboxylated into acetyl coenzyme A (acetyl-CoA).…”

Section: Yeasts At Work: Nectar Fermentation and Voc Productionmentioning

confidence: 99%

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Nature’s Most Fruitful Threesome: The Relationship between Yeasts, Insects, and Angiosperms

Fenner

Scapini

Diniz

et al. 2022

JoF

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The importance of insects for angiosperm pollination is widely recognized. In fact, approximately 90% of all plant species benefit from animal-mediated pollination. However, only recently, a third part player in this story has been properly acknowledged. Microorganisms inhabiting floral nectar, among which yeasts have a prominent role, can ferment glucose, fructose, sucrose, and/or other carbon sources in this habitat. As a result of their metabolism, nectar yeasts produce diverse volatile organic compounds (VOCs) and other valuable metabolites. Notably, some VOCs of yeast origin can influence insects’ foraging behavior, e.g., by attracting them to flowers (although repelling effects have also been reported). Moreover, when insects feed on nectar, they also ingest yeast cells, which provide them with nutrients and protect them from pathogenic microorganisms. In return, insects serve yeasts as transportation and a safer habitat during winter when floral nectar is absent. From the plant’s point of view, the result is flowers being pollinated. From humanity’s perspective, this ecological relationship may also be highly profitable. Therefore, prospecting nectar-inhabiting yeasts for VOC production is of major biotechnological interest. Substances such as acetaldehyde, ethyl acetate, ethyl butyrate, and isobutanol have been reported in yeast volatomes, and they account for a global market of approximately USD 15 billion. In this scenario, the present review addresses the ecological, environmental, and biotechnological outlooks of this three-party mutualism, aiming to encourage researchers worldwide to dig into this field.

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“…where PAi is the abundance of protein i, N is the total number of proteins in the measured proteome, and ∑ 𝑓 𝑃𝑖 𝑁 𝑖=1 = 1. Mass fraction of a pathway proteome, fPath, that contains M proteins is determined by (82):…”

mentioning

confidence: 99%

Proteomes Reveal Metabolic Capabilities ofYarrowia lipolyticafor Biological Upcycling of Polyethylene into High-Value Chemicals

Walker

Mortensen

Poudel

et al. 2023

Preprint

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Polyolefins derived from plastic wastes are recalcitrant for biological upcycling. However, chemical depolymerization of polyolefins can generate depolymerized plastic (DP) oil comprising of a complex mixture of saturated, unsaturated, even and odd hydrocarbons suitable for biological conversion. While DP oil contains a rich carbon and energy source, it is inhibitory to cells. Understanding and harnessing robust metabolic capabilities of microorganisms to upcycle the hydrocarbons in DP oil, both naturally and unnaturally occurring, into high-value chemicals are limited. Here, we discovered that an oleaginous yeastYarrowia lipolyticaundergoing short-term adaptation to DP oil robustly utilized a wide range of hydrocarbons for cell growth and production of citric acid and neutral lipids. When growing on hydrocarbons,Y. lipolyticapartitioned into planktonic and oil-bound cells with each exhibiting distinct proteomes and amino acid distributions invested into establishing these proteomes. Significant proteome reallocation towards energy and lipid metabolism, belonging to two of the 23 KOG (Eukaryotic Orthologous Groups) classes C and I, enabled robust growth ofY. lipolyticaon hydrocarbons, with n-hexadecane as the preferential substrate. This investment was even higher for growth on DP oil where both the KOG classes C and I were the top two, and many associated proteins and pathways were expressed and upregulated including the hydrocarbon degradation pathway, Krebs cycle, glyoxylate shunt and, unexpectedly, propionate metabolism. However, a reduction in proteome allocation for protein biosynthesis, at the expense of the observed increase towards energy and lipid metabolisms, might have caused the inhibitory effect of DP oil on cell growth.

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Proteome reallocation enables the selective de novo biosynthesis of non-linear, branched-chain acetate esters

Cited by 8 publications

References 56 publications

Engineering a Synthetic Escherichia coli Coculture for Compartmentalized de novo Biosynthesis of Isobutyl Butyrate from Mixed Sugars

Engineering a Synthetic Escherichia coli Coculture for Compartmentalized de novo Biosynthesis of Isobutyl Butyrate from Mixed Sugars

Nature’s Most Fruitful Threesome: The Relationship between Yeasts, Insects, and Angiosperms

Proteomes Reveal Metabolic Capabilities ofYarrowia lipolyticafor Biological Upcycling of Polyethylene into High-Value Chemicals

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