Arbutin, a glycoside, is derived from the leaves of several plants, including wheat, pear, and bearberry plants, and has a significant role in the treatment of melanoma, cystitis, and cough. Here, we aimed to modify Yarrowia lipolytica to produce arbutin. To construct the arbutin synthetic pathway in Y. lipolytica, three genes (chorismate pyruvate-lyase (UbiC), 4-hydroxybenzoate 1-hydroxylase (MNX1), and hydroquinone glucosyltransferase (AS)) were codon-optimized and heterologously expressed. To maximize arbutin production, seven arbutin-biosynthesis molecular targets were overexpressed, and we found that the individual strengthening of DHS1 and DHS2 led to an 8.9- and 7.8-fold improvement in arbutin yield, respectively. Through optimization, a maximum arbutin titer of 8.6 ± 0.7 g/L was achieved using the finally engineered strain, po1f-At09. Overall, this is the first report of heterologous arbutin synthesis in Y. lipolytica at a high titer. Furthermore, this work opens a possibility for the overproduction of shikimate pathway derivatives in Y. lipolytica.

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Biosensors regulated by specific substrates are needed to develop genetic tools to meet the needs of engineering microbial cell factories. Here, a xylose-inducible biosensor (xylbiosensor), comprising the Escherichia coli activation factor XylR, fusion activation domain (AD) VPRH, and a hybrid promoter with operator xylO, was established in Yarrowia lipolytica. The addition of xylose to an engineered Y. lipolytica strain harboring the xylbiosensor could trigger significant transcriptional activation of target genes, such as mcherry and the xylose utilization gene. Furthermore, a novel promoter Pleu-Pxo-Ptef was developed to construct a bidirectional expression system. The xylbiosensor showed good portability in Saccharomyces cerevisiae, suggesting its potential value in other eukaryotic cells. This study is the first to construct a “turn-on” xylbiosensor induced by xylose addition based on a prokaryotic activator XylR and eukaryotic universal AD. The xylbiosensor exhibits potential in pathway engineering for xylose utilization and xylose-derived product biosynthesis in yeast.

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