The wealth of bio-based building blocks produced by engineered microorganisms seldom include halogen atoms. Muconate is a platform chemical with a number of industrial applications that could be broadened by introducing fluorine atoms to tune its physicochemical properties. The soil bacterium Pseudomonas putida naturally assimilates benzoate via the ortho-cleavage pathway with cis,cis-muconate as intermediate. Here, we harnessed the native enzymatic machinery (encoded within the ben and cat gene clusters) to provide catalytic access to 2-fluoro-cis,cis-muconate (2-FMA) from fluorinated benzoates. The reactions in this pathway are highly imbalanced, leading to accumulation of toxic intermediates and limited substrate conversion. By disentangling regulatory patterns of ben and cat in response to fluorinated effectors, metabolic activities were adjusted to favor 2-FMA biosynthesis. After implementing this combinatorial approach, engineered P. putida converted 3-fluorobenzoate to 2-FMA at the maximum theoretical yield. Hence, this study illustrates how synthetic biology can expand the diversity of nature's biochemical catalysis.

由工程微生物产生的大量生物基构建块很少包含卤素原子。Muconate 是一种具有多种工业应用的平台化学品，可以通过引入氟原子来调整其物理化学性质来扩大应用范围。土壤细菌恶臭假单胞菌通过以顺式、顺式粘康酸为中间体的邻位裂解途径自然同化苯甲酸。在这里，我们利用天然酶促机制（编码在ben和cat基因簇中）提供对 2-氟顺式、顺式-来自氟化苯甲酸盐的粘康酸盐 (2-FMA)。该途径中的反应高度不平衡，导致有毒中间体的积累和底物转化受限。通过解开ben和cat响应氟化效应物的调节模式，调整代谢活动以有利于 2-FMA 生物合成。在实施这种组合方法后，设计了P。恶臭以最大理论产率将 3-氟苯甲酸酯转化为 2-FMA。因此，这项研究说明了合成生物学如何扩展自然界生化催化的多样性。