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Linkage-Specific Detection and Metabolism of Human Milk Oligosaccharides in Escherichia coli
Cell Chemical Biology ( IF 6.6 ) Pub Date : 2018-07-12 , DOI: 10.1016/j.chembiol.2018.06.002
Fatima Enam , Thomas J. Mansell

Human milk oligosaccharides (HMOs) are important prebiotic complex carbohydrates with demonstrated beneficial effects on the microbiota of neonates. However, optimization of their biotechnological synthesis is limited by the relatively low throughput of monosaccharide and linkage analysis. To enable high-throughput screening of HMO structures, we constructed a whole-cell biosensor that uses heterologous expression of glycosidases to generate linkage-specific, quantitative fluorescent readout for a range of HMOs at detection limits down to 20 μM in approximately 6 hr. We also demonstrate the use of this system for orthogonal control of growth rate or protein expression of particular strains in mixed populations. This work enables rapid non-chromatographic linkage analysis and lays the groundwork for the application of directed evolution to biosynthesis of complex carbohydrates as well as the prebiotic manipulation of population dynamics in natural and engineered microbial communities.

中文翻译:

大肠杆菌中人乳寡糖的连锁特异性检测和代谢

人乳低聚糖(HMO)是重要的益生元复合碳水化合物,对新生儿的微生物群具有有益的作用。但是,单糖和连接分析的相对较低的通量限制了其生物技术合成的优化。为了能够对HMO结构进行高通量筛选,我们构建了一个全细胞生物传感器,该传感器使用糖苷酶的异源表达在约6小时内检测到最低20μM的HMO范围内产生连锁特异性,定量荧光读数。我们还证明了该系统用于混合种群中特定菌株的生长速率或蛋白质表达的正交控制。
更新日期:2018-10-19
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