Engineering a Synthetic Pathway for Tyrosol Synthesis in Escherichia coli.

Tyrosol is an aromatic compound with great value that is widely used in the food and pharmaceutical industry. In this study, we reported a synthetic pathway for converting p-coumaric acid (p-CA) into tyrosol in Escherichia coli. We found that the enzyme cascade comprising ferulic acid decarboxylase (FDC1) from Saccharomyces cerevisiae, styrene monooxygenase (SMO), styrene oxide isomerase (SOI) from Pseudomonas putida, and phenylacetaldehyde reductase (PAR) from Solanum lycopersicum could efficiently synthesize tyrosol from p-CA with a conversion rate over 90%. To further expand the range of substrates, we also introduced tyrosine ammonia-lyase (TAL) from Flavobacterium johnsoniae to connect the synthetic pathway with the endogenous l-tyrosine metabolism. We found that tyrosol could be efficiently produced from glycerol, reaching 545.51 mg/L tyrosol in a tyrosine-overproducing strain under shake flasks. In summary, we have established alternative routes for tyrosol synthesis from p-CA (a potential lignin-derived biomass), glucose, and glycerol.

One-Pot Bioconversion of Lignin-Derived Substrates into Gallic Acid.

Lignin is regarded as the most abundant renewable aromatic compound on earth. In this study, we established Escherichia coli-based whole-cell biocatalytic systems to efficiently convert two lignin-derived substrates (ferulic acid and p-coumaric acid) to gallic acid. For the synthesis of gallic acid from ferulic acid, we used the recombinant E. coli expressing feruloyl-CoA synthetase and enoyl-CoA hydratase/aldolase from Pseudomonas putida, aldehyde dehydrogenase (HFD1) from Saccharomyces cerevisiae, vanillic acid O-demethylase (VanAB) from P. putida, and a mutant version of p-hydroxybenzoate hydroxylase (PobAY385F) from P. putida. Under the fed-batch mode, 19.57 mM gallic acid was obtained from 20 mM ferulic acid with a conversion rate of 97.9%. To achieve gallic acid synthesis from p-coumaric acid, we replaced VanAB with the two-component flavin-dependent monooxygenase (HpaBC) from E. coli. Under optimal conditions, 20 mM p-coumaric acid afforded the production of 19.96 mM gallic acid with near 100% conversion. To the best of our knowledge, our work represented the first study to develop E. coli-based whole-cell biocatalysts for the eco-friendly synthesis of gallic acid from lignin-derived renewable feedstocks.

Utilization of a styrene-derived pathway for 2-phenylethanol production in budding yeast.

2-Phenylethanol (2-PE) is an important flavor ingredient and is widely applied in the fields of food, cosmetics, and pharmaceuticals. Despite that Saccharomyces cerevisiae has the ability to naturally synthesize 2-PE via the Ehrlich pathway, de novo synthesis of 2-PE in high titer still remains a huge challenge. In this study, a non-native styrene degradation pathway was introduced into S. cerevisiae, which represents the first time to demonstrate the functional expression of "styrene-derived" 2-PE synthesis in yeast. Using a host strain engineered with L-phenylalanine (L-Phe) overproduction, the heterologous 2-PE pathway coupled with endogenous Ehrlich pathway produced 233 mg/L 2-PE under shake flasks. Additionally, we further engineered the permease transporters to improve the intracellular L-Phe availability, and further improved the 2-PE titer to 680 mg/L. Taken together, our work represents one of the pioneering reports to explore "styrene-derived" pathway in S. cerevisiae. The synthetic yeast described here might be used as a platform for the future development of next-generation high-yielding 2-PE yeast strains.Key Points• A styrene-derived pathway was established in yeast for 2-phenylethanol productions; membrane-associated styrene oxide isomerase was functional in yeast.• Transporter engineering to improve the L-phenylalanine importation with enhanced 2-phenylethanol productions.

The function of KptA/Tpt1 gene - a minor review.

Rapid response of uni- and multicellular organisms to environmental changes and their own growth is achieved through a series of molecular mechanisms, often involving modification of macromolecules, including nucleic acids, proteins and lipids. The ADP-ribosylation process has ability to modify these different macromolecules in cells, and is closely related to the biological processes, such as DNA replication, transcription, signal transduction, cell division, stress, microbial aging and pathogenesis. In addition, tRNA plays an essential role in the regulation of gene expression, as effector molecules, no-load tRNA affects the overall gene expression level of cells under some nutritional stress. KptA/Tpt1 is an essential phosphotransferase in the process of pre-tRNA splicing, releasing mature tRNA and participating in ADP-ribose. The objective of this review is concluding the gene structure, the evolution history and the function of KptA/Tpt1 from prokaryote to eukaryote organisms. At the same time, the results of promoter elements analysis were also shown in the present study. Moreover, the problems in the function of KptA/Tpt1 that have not been clarified at the present time are summarised, and some suggestions to solve those problems are given. This review presents no only a summary of clear function of KptA/Tpt1 in the process of tRNA splicing and ADP-ribosylation of organisms, but also gives some proposals to clarify unclear problems of it in the future.