Kirtz Marko, Klebensberger Janosch, Otte Konrad B, Richter Sven M, Hauer Bernhard
Institute of Technical Biochemistry, University of Stuttgart, Allmandring 31, 70569 Stuttgart, Germany.
Institute of Technical Biochemistry, University of Stuttgart, Allmandring 31, 70569 Stuttgart, Germany.
J Biotechnol. 2016 Jul 20;230:30-3. doi: 10.1016/j.jbiotec.2016.05.017. Epub 2016 May 13.
The present proof-of-concept study reports the construction of a whole-cell biocatalyst for the de novo production of ω-hydroxy octanoic acid. This was achieved by hijacking the natural fatty acid cycle and subsequent hydroxylation using a specific monooxygenase without the need for the additional feed of alkene-like precursors. For this, we used the model organism Escherichia coli and increased primarily the release of the octanoic acid precursors by overexpressing the plant thioesterase FatB2 from Cuphea hookeriana in a β-oxidation deficient strain, which lead to the production of 2.32mM (8.38mggcww(-1)) octanoic acid in 24h. In order to produce the corresponding ω-hydroxy derivative, we additionally expressed the engineered self-sufficient monooxygenase fusion protein CYP153AMaq(G307A)-CPRBM3 within the octanoic acid producing strain. With this, we finally produced 234μM (0.95mggcww(-1)) ω-hydroxy octanoic acid in a 20h fed-batch set-up.
本概念验证研究报告了一种用于从头生产ω-羟基辛酸的全细胞生物催化剂的构建。这是通过利用天然脂肪酸循环并随后使用特定的单加氧酶进行羟基化来实现的,无需额外添加类似烯烃的前体。为此,我们使用了模式生物大肠杆菌,并通过在β-氧化缺陷菌株中过表达来自霍氏萼距花的植物硫酯酶FatB2,主要增加了辛酸前体的释放,这导致在24小时内产生了2.32 mM(8.38 mg g cw w(-1))的辛酸。为了生产相应的ω-羟基衍生物,我们在产生辛酸的菌株中额外表达了工程化的自给自足单加氧酶融合蛋白CYP153AMaq(G307A)-CPRBM3。通过这种方式,我们最终在20小时的补料分批培养设置中生产了234 μM(0.95 mg g cw w(-1))的ω-羟基辛酸。