ACS Synthetic Biology最新文献

The use of synthetic antigen-presenting cells to activate and expand engineered T cells for the treatment of cancers typically results in therapies that are suboptimal in effectiveness and durability. Here we describe a high-throughput microfluidic system for the fabrication of synthetic cells mimicking the viscoelastic and T-cell-activation properties of antigen-presenting cells. Compared with rigid or elastic microspheres, the synthetic viscoelastic T-cell-activating cells (SynVACs) led to substantial enhancements in the expansion of human CD8⁺ T cells and to the suppression of the formation of regulatory T cells. Notably, activating and expanding chimaeric antigen receptor (CAR) T cells with SynVACs led to a CAR-transduction efficiency of approximately 90% and to substantial increases in T memory stem cells. The engineered CAR T cells eliminated tumour cells in a mouse model of human lymphoma, suppressed tumour growth in mice with human ovarian cancer xenografts, persisted for longer periods and reduced tumour-recurrence risk. Our findings underscore the crucial roles of viscoelasticity in T-cell engineering and highlight the utility of SynVACs in cancer therapy.

Salicylate plays a pivotal role as a pharmaceutical intermediate in drugs, such as aspirin and lamivudine. The low catalytic efficiency of key enzymes and the inherent toxicity of salicylates to cells pose significant challenges to large-scale microbial production. In this study, we introduced the salicylate synthase Irp9 into an l-phenylalanine-producing Escherichia coli, constructing the shortest salicylate biosynthetic pathway. Subsequent protein engineering increased the catalytic efficiency of Irp9 by 33.5%. Furthermore, by integrating adaptive evolution with transcriptome analysis, we elucidated the crucial mechanism of efflux proteins in salicylate tolerance. The elucidation of this mechanism guided us in the targeted modification of these transport proteins, achieving a reported maximum level of 3.72 g/L of salicylate in a shake flask. This study highlights the importance of efflux proteins for enhancing the productivity of microbial cell factories in salicylate production, which also holds potential for application in the green synthesis of other phenolic acids.

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The declining availability of cheap fossil-based resources has sparked growing interest in the sustainable biosynthesis of organic acids. l-Malic acid, a crucial four-carbon dicarboxylic acid, finds extensive applications in the food, chemical, and pharmaceutical industries. Synthetic biology and metabolic engineering have enabled the efficient microbial production of l-malic acid, albeit not in Yarrowia lipolytica, an important industrial microorganism. The present study aimed to explore the potential of this fungal species for the production of l-malic acid. First, endogenous biosynthetic genes and heterologous transporter genes were overexpressed in Y. lipolytica to identify bottlenecks in the l-malic acid biosynthesis pathway grown on glycerol. Second, overexpression of isocitrate lyase, malate synthase, and malate dehydrogenase in the glyoxylate cycle pathway and introduction of a malate transporter from Schizosaccharomyces pombe significantly boosted l-malic acid production, which reached 27.0 g/L. A subsequent increase to 37.0 g/L was attained through shake flask medium optimization. Third, adaptive laboratory evolution allowed the engineered strain Po1g-CEE2+Sp to tolerate a lower pH and to accumulate a higher amount of l-malic acid (56.0 g/L). Finally, when scaling up to a 5 L bioreactor, a titer of 112.5 g/L was attained. In conclusion, this study demonstrates for the first time the successful production of l-malic acid in Y. lipolytica by combining metabolic engineering and laboratory evolution, paving the way for large-scale sustainable biosynthesis of this and other organic acids.

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