Sakuranetin is a plant-natural product, which has increasingly been utilized in cosmetic and pharmaceutical industries for its extensive anti-inflammatory, anti-tumor, and immunomodulatory effects. Sakuranetin was mostly produced via extraction technology from plants, which is limited to natural conditions and biomass supply. In this study, a novel strategy to produce sakuranetin via de novo synthesis from glucose by engineering S. cerevisiae was introduced . After a series of heterogenous genes integration, a biosynthetic pathway of sakuranetin from glucose was successfully constructed in S. cerevisiae which sakuranetin yield reached only 4.28 mg/L. Then, a multi-module metabolic engineering strategy was applied for improving sakuranetin yield in S. cerevisiae: (1) adjusting the copy number of sakuranetin synthesis genes; (2) removing the rate-limiting factor of aromatic amino pathway and optimizing the synthetic pathway of aromatic amino acids to enhance the supply of carbon flux for sakuranetin; (3) introducing acetyl-CoA carboxylase mutants ACC1S659A, S1157A, and knocking-out YPL062W to strengthen the supply of malonyl-CoA which is another synthetic precursor of sakuranetin. The resultant mutant S. cerevisiae exhibited a more than 10-fold increase of sakuranetin titer (50.62 mg/L) in shaking flasks. Furthermore, the sakuranetin titer increased to 158.65 mg/L in a 1-L bioreactor, which is the highest sakuranetin titer among all publications reported yield of the engineered microbial cell.

Dibutyl phthalate (DBP) is an environmental pollutant that can threaten human health. The strain Arthrobacter sp. ZJUTW, isolated from the sludge of river of Hangzhou city, can efficiently degrade DBP. Its genomic and transcriptomic differences when cultivated with DBP and with glucose revealed specific DBP metabolic pathways in the ZJUTW strain. The degrading gene clusters distribute separately on a circular chromosome and a plasmid pQL1. Genes related to the initial steps of DBP degradation from DBP to phthalic acid (PA), the pehA gene and pht gene cluster, are located on the plasmid pQL1. While pca gene cluster related to the transforming of protocatechuic acid (PCA) to acetyl-CoA, is located on the chromosome. After homologous alignment analysis with the reported gene clusters, we found that there were a series of double copies of homologous genes in pht and pca gene clusters that contribute to the efficient degradation of DBP by ZJUTW. In addition, transcriptomic analysis showed a synergistic effect between pht and pca clusters, which also favor ZJUTW allowing it to efficiently degrade DBP. Combined genomic and transcriptomic analyses affords the complete DBP metabolic pathway in Arthrobacter sp. ZJUTW that is different from that of reported other Arthrobacter strains. After necessary modification based on its metabolic characteristics, Arthrobacter sp. ZJUTW or its mutants might represent promising candidates for use in the bioremediation of DBP pollution.