Loeschcke Anita, Markert Annette, Wilhelm Susanne, Wirtz Astrid, Rosenau Frank, Jaeger Karl-Erich, Drepper Thomas
Institute of Molecular Enzyme Technology, Heinrich-Heine-University Düsseldorf, Research Center Jülich, Jülich, Germany.
ACS Synth Biol. 2013 Jan 18;2(1):22-33. doi: 10.1021/sb3000657. Epub 2012 Oct 17.
Secondary metabolites represent a virtually inexhaustible source of natural molecules exhibiting a high potential as pharmaceuticals or chemical building blocks. To gain broad access to these compounds, sophisticated expression systems are needed that facilitate the transfer and expression of large chromosomal regions, whose genes encode complex metabolic pathways. Here, we report on the development of the novel system for the transfer and expression of biosynthetic pathways (TREX), which comprises all functional elements necessary for the delivery and concerted expression of clustered pathway genes in different bacteria. TREX employs (i) conjugation for DNA transfer, (ii) randomized transposition for its chromosomal insertion, and (iii) T7 RNA polymerase for unimpeded bidirectional gene expression. The applicability of the TREX system was demonstrated by establishing the biosynthetic pathways of two pigmented secondary metabolites, zeaxanthin and prodigiosin, in bacteria with different metabolic capacities. Thus, TREX represents a valuable tool for accessing natural products by allowing comparative expression studies with clustered genes.
次生代谢产物是天然分子的一个几乎取之不尽的来源，作为药物或化学构建模块具有很高的潜力。为了广泛获取这些化合物，需要复杂的表达系统来促进大染色体区域的转移和表达，这些区域的基因编码复杂的代谢途径。在此，我们报告了一种用于生物合成途径转移和表达的新型系统（TREX）的开发，该系统包含在不同细菌中传递和协同表达成簇途径基因所需的所有功能元件。TREX采用（i）接合进行DNA转移，（ii）随机转座进行染色体插入，以及（iii）T7 RNA聚合酶进行不受阻碍的双向基因表达。通过在具有不同代谢能力的细菌中建立两种色素次生代谢产物玉米黄质和灵菌红素的生物合成途径，证明了TREX系统的适用性。因此，TREX通过允许对成簇基因进行比较表达研究，代表了一种获取天然产物的有价值工具。
