Droplet-based microfluidic high-throughput screening of heterologous enzymes secreted by the yeast Yarrowia lipolytica

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• 作者：Thomas Beneyton ; Stéphane Thomas ; Andrew D. Griffiths…
• 关键词：High ; throughput enzymatic screening ; Droplet ; based microfluidics ; Heterologous expression ; Yarrowia lipolytica ; Recombinant fungal protein
• 刊名：Microbial Cell Factories
• 出版年：2017
• 出版时间：December 2017
• 年：2017
• 卷：16
• 期：1
• 全文大小：2058KB
• 刊物类别：Chemistry and Materials Science
• 刊物主题：Applied Microbiology; Biotechnology; Microbiology; Microbial Genetics and Genomics; Enzymology; Genetic Engineering;
• 出版者：BioMed Central
• ISSN：1475-2859
• 卷排序：16

文摘

BackgroundDroplet-based microfluidics is becoming an increasingly attractive alternative to microtiter plate techniques for enzymatic high-throughput screening (HTS), especially for exploring large diversities with lower time and cost footprint. In this case, the assayed enzyme has to be accessible to the substrate within the water-in-oil droplet by being ideally extracellular or displayed at the cell surface. However, most of the enzymes screened to date are expressed within the cytoplasm of Escherichia coli cells, which means that a lysis step must take place inside the droplets for enzyme activity to be assayed. Here, we take advantage of the excellent secretion abilities of the yeast Yarrowia lipolytica to describe a highly efficient expression system particularly suitable for the droplet-based microfluidic HTS.