Integrated pathway engineering and transcriptome analysis for improved astaxanthin biosynthesis in Yarrowia lipolytica

Astaxanthin is a high value carotenoid with a broad range of commercial applications due to its superior antioxidant properties.In this study,β-carotene-producing Yarrowia lipolytica XK17 constructed in the lab was employed for astaxanthin biosynthesis.The catalytic effects ofβ-carotene ketolase CrtW andβ-carotene hydroxylase CrtZ from various species were investigated.The PspCrtW from Paracoccus sp.and HpCrtZ^(#) from Haematococcus pluvialis were confirmed to be the best combination in convertingβ-carotene.Several key bottlenecks in biomass and astaxanthin biosynthesis were effectively eliminated by optimizing the expression of the above enzymes and restoring uracil/leucine biosynthesis.In addition,the effects of astaxanthin biosynthesis on cell metabolism were investigated by integrated analysis of pathway modification and transcriptome information.After further optimization,strain DN30 was able to synthesize up to 730.3 mg/L astaxanthin in laboratory 5-L fermenter.This study provides a good metabolic strategy and a sustainable development platform for high-value carotenoid production.

Recent advances in microbial production of phenolic compounds

Phenolic compounds(PCs)are a group of compounds with various applications in nutraceutical,pharmaceutical and cosmetic industries.Their supply by plant extraction and chemical synthesis is often limited by low yield and high cost.Microbial production represents as a promising alternative for efficient and sustainable production of PCs.In this review,we summarize recent advances in this field,which include enzyme mining and engineering to construct artificial pathways,balance of enzyme expression to improve pathway efficiency,coculture engineering to alleviate metabolic burden and side-reactions,and the use of genetic circuits for dynamic regulation and high throughput screening.Finally,current challenges and future perspectives for efficient production of PCs are also discussed.

Design and thermodynamic analysis of a pathway enabling anaerobic production of poly-3-hydroxybutyrate in Escherichia coli

Utilizing anaerobic metabolisms for the production of biotechnologically relevant products presents potential advantages,such as increased yields and reduced energy dissipation.However,lower energy dissipation may indicate that certain reactions are operating closer to their thermodynamic equilibrium.While stoichiometric analyses and genetic modifications are frequently employed in metabolic engineering,the use of thermodynamic tools to evaluate the feasibility of planned interventions is less documented.In this study,we propose a novel metabolic engineering strategy to achieve an efficient anaerobic production of poly-(R)-3-hydroxybutyrate(PHB)in the model organism Escherichia coli.Our approach involves re-routing of two-thirds of the glycolytic flux through non-oxidative glycolysis and coupling PHB synthesis with NADH re-oxidation.We complemented our stoichiometric analysis with various thermodynamic approaches to assess the feasibility and the bottlenecks in the proposed engineered pathway.According to our calculations,the main thermodynamic bottleneck are the reactions catalyzed by the acetoacetyl-CoAβ-ketothiolase(EC 2.3.1.9)and the acetoacetyl-CoA reductase(EC 1.1.1.36).Furthermore,we calculated thermodynamically consistent sets of kinetic parameters to determine the enzyme amounts required for sustaining the conversion fluxes.In the case of the engineered conversion route,the protein pool necessary to sustain the desired fluxes could account for 20%of the whole cell dry weight.

Engineering bacteria for enhanced polyhydroxyalkanoates(PHA)biosynthesis

Polyhydroxyalkanoates(PHA)have been produced by some bacteria as bioplastics for many years.Yet their commercialization is still on the way.A few issues are related to the difficulty of PHA commercialization:namely,high cost and instabilities on molecular weights(Mw)and structures,thus instability on thermo-mechanical properties.The high cost is the result of complicated bioprocessing associated with sterilization,low conversion of carbon substrates to PHA products,and slow growth of microorganisms as well as difficulty of downstream separation.Future engineering on PHA producing microorganisms should be focused on contamination resistant bacteria especially extremophiles,developments of engineering approaches for the extremophiles,increase on carbon substrates to PHA conversion and controlling Mw of PHA.The concept proof studies could still be conducted on E.coli or Pseudomonas spp.that are easily used for molecular manipulations.In this review,we will use E.coli and halophiles as examples to show how to engineer bacteria for enhanced PHA biosynthesis and for increasing PHA competitiveness.

Advances in engineering methylotrophic yeast for biosynthesis of valuable chemicals from methanol

Methylotrophic yeasts and bacteria, which can use methanol as carbon and energy source, have beenwildly used as microbial cell factories for biomanufacturing. Due to their robustness in industrial harshconditions, methylotrophic yeasts such as Pichia pastoris have been explored as a cell factory forproduction of proteins and high-value chemicals. Methanol utilization pathway （MUT） is highlyregulated for efficient methanol utilization, and the downstream pathways need extensively constructedand optimized toward target metabolite biosynthesis. Here, we present an overview of methanolmetabolism and regulation in methylotrophic yeasts, among which we focus on the regulation of keygenes involved in methanol metabolism. Besides, the recent progresses in construction and optimizationof downstream biosynthetic pathways for production of high value chemicals, such as polyketides, fattyacids and isoprenoids, are further summarized. Finally, we discuss the current challenges and feasiblestrategies toward constructing efficient methylotrophic cell factories may promote wide applications inthe future.

Issues and strategies of cathode materials for mild aqueous static zinc-ion batteries

Researchers prefer mild aqueous static zinc-ion batteries(ASZIBs)for their distinct benefits of excellent safety,abundant zinc resources,low cost,and high energy density.However,at the moment there are some issues with the cathode materials of mild ASZIBs,including dissolution,by-products,poor conductivity,and a contentious energy storage system.Consequently,there are numerous difficulties in the development of high-performance mild ASZIBs cathode materials.This overview examines the mechanisms for storing energy and the de-velopments in inorganic,organic,and other novel cathode materials that have emerged in recent years.At the same time,three solutions—structural engineering,interface engineering,and reaction pathway engineering—as well as the difficulties now faced by the cathode materials of mild ASZIBs are forcefully introduced.Finally,a prospect is made regarding the evolution of cathode materials in the future.