CAS Key Laboratory of Synthetic Biology, CAS Center for Excellence in Molecular Plant Sciences, Institute of Plant Physiology and Ecology , Chinese Academy of Sciences , Shanghai 200032 , China.
University of Chinese Academy of Sciences , Beijing 100039 , China.
ACS Chem Biol. 2018 Aug 17;13(8):1944-1949. doi: 10.1021/acschembio.8b00285. Epub 2018 Jun 12.
Functional discovery and characterization of the target enzymes responsible for the biosynthesis pathway coded for the genes is ongoing, and the unknown functional diversity of this class of enzymes has been revealed by genome sequencing. Commonly, it is feasible in annotating of biosynthetic genes of prokaryotes due to the existence of gene clusters of secondary metabolites. However, in eukaryotes, the biosynthetic genes are not compactly clustered in the way of prokaryotes. Hence, it remains challenging to identify the biosynthetic pathways of newly discovered natural products in plants. Steviol glycosides are one class of natural sweeteners found in high abundance in the herb Stevia rebaudiana. Here, we applied the chemoproteomic strategy for the proteomic profiling of the biosynthetic enzymes of steviol glycosides in Stevia rebaudiana. We not only identified a steviol-catalyzing UDP-glycosyltransferase (UGT) UGT73E1 involved in steviol glycoside biosynthesis but also built up a probe-based platform for the screening of potential substrates of functional uncharacterized UGT rapidly. This approach would be a complementary tool in mining novel synthetic parts for assembling of synthetic biological systems for the biosynthesis of other complex natural products.
功能发现和鉴定负责基因编码生物合成途径的靶酶的工作正在进行中，基因组测序揭示了这一类酶的未知功能多样性。通常，由于次级代谢物基因簇的存在，在原核生物的生物合成基因注释中是可行的。然而，在真核生物中，生物合成基因不像原核生物那样紧密聚集。因此，识别植物中新发现的天然产物的生物合成途径仍然具有挑战性。甜菊醇糖苷是在甜叶菊中大量存在的一类天然甜味剂。在这里，我们应用化学生物组学策略对甜菊醇糖苷的生物合成酶进行蛋白质组学分析。我们不仅鉴定了参与甜菊醇糖苷生物合成的甜菊醇催化 UDP-糖基转移酶（UGT）UGT73E1，还建立了一个基于探针的平台，用于快速筛选功能未知的 UGT 的潜在底物。这种方法将是挖掘用于组装合成生物系统以合成其他复杂天然产物的新型合成部件的互补工具。
