Chou Alexander, Lee Seung Hwan, Zhu Fayin, Clomburg James M, Gonzalez Ramon
Department of Chemical, Biological and Materials Engineering, University of South Florida, Tampa, FL, USA.
Nat Metab. 2021 Oct;3(10):1385-1399. doi: 10.1038/s42255-021-00453-0. Epub 2021 Oct 21.
Metabolic engineering often entails concurrent engineering of substrate utilization, central metabolism and product synthesis pathways, inevitably creating interdependency with native metabolism. Here we report an alternative approach using synthetic pathways for C1 bioconversion that generate multicarbon products directly from C1 units and hence are orthogonal to the host metabolic network. The engineered pathways are based on formyl-CoA elongation (FORCE) reactions catalysed by the enzyme 2-hydroxyacyl-CoA lyase. We use thermodynamic and stoichiometric analyses to evaluate FORCE pathway variants, including aldose elongation, α-reduction and aldehyde elongation. Promising variants were prototyped in vitro and in vivo using the non-methylotrophic bacterium Escherichia coli. We demonstrate the conversion of formate, formaldehyde and methanol into various products including glycolate, ethylene glycol, ethanol and glycerate. FORCE pathways also have the potential to be integrated with the host metabolism for synthetic methylotrophy by the production of native growth substrates as demonstrated in a two-strain co-culture system.
代谢工程通常需要对底物利用、中心代谢和产物合成途径进行同步工程改造，这不可避免地会与天然代谢产生相互依赖性。在此，我们报告了一种利用合成途径进行C1生物转化的替代方法，该方法可直接从C1单元生成多碳产物，因此与宿主代谢网络正交。工程化途径基于由2-羟基酰基辅酶A裂解酶催化的甲酰基辅酶A延伸（FORCE）反应。我们使用热力学和化学计量分析来评估FORCE途径变体，包括醛糖延伸、α-还原和醛延伸。使用非甲基营养型细菌大肠杆菌在体外和体内对有前景的变体进行了原型设计。我们展示了将甲酸盐、甲醛和甲醇转化为各种产物，包括乙醇酸、乙二醇、乙醇和甘油酸。如在双菌株共培养系统中所示，FORCE途径还具有通过产生天然生长底物与宿主代谢整合以实现合成甲基营养的潜力。
