Escherichia coli Deletion Mutants Illuminate Trade-Offs between Growth Rate and Flux through a Foreign Anabolic Pathway

Metabolic engineers strive to improve the production yields of microbial fermentations, sometimes by mutating the genomes of production strains. Some mutations are detrimental to the health of the organism, so a quantitative and mechanistic understanding of the trade-offs could inform better designs. We employed the bacterial luciferase operon (luxABCDE), which uses ubiquitous energetic cofactors (NADPH, ATP, FMNH2, acetyl-CoA) from the host cell, as a proxy for a novel anabolic pathway. The strains in the Escherichia coli Keio collection, each of which contains a single deletion of a non-essential gene, represent mutational choices that an engineer might make to optimize fermentation yields. The Keio strains and the parental BW25113 strain were transformed with a luxABCDE expression vector. Each transformant was propagated in defined M9 medium at 37°C for 48 hours; the cell density (optical density at 600 nanometers, OD600) and luminescence were measured every 30 minutes. The trade-offs were visualized by plotting the maximum growth rate and luminescence/OD600 of each transformant across a “production possibility frontier”. Our results show that some loss-of-function mutations enhance growth in vitro or light production, but that improvement in one trait generally comes at the expense of the other.

代谢工程研究者致力于提升微生物发酵的产物得率，有时会通过诱变生产菌株的基因组来实现目标。部分突变会对菌体生长产生不利影响，因此对这类性状权衡的定量与机制层面解析，可为更优化的工程设计提供依据。本研究选用细菌荧光素酶操纵子（luxABCDE）作为新型合成代谢途径的替代模型——该操纵子可利用宿主细胞内普遍存在的能量辅因子（NADPH、ATP、FMNH₂、乙酰辅酶A（acetyl-CoA））。本研究使用的大肠杆菌Keio敲除文库（Escherichia coli Keio Collection）中的每一株菌株均携带单个非必需基因的敲除，对应了工程菌株优化发酵得率时可能采取的诱变策略。将Keio文库菌株与野生型亲本菌株BW25113均转化带有luxABCDE的表达载体。每一株转化子均在37℃下于化学成分明确的M9培养基中培养48小时；每30分钟测定一次菌体密度（600纳米处光密度值，OD₆₀₀）与发光强度。研究通过绘制每株转化子的最大生长速率与发光强度/OD₆₀₀比值的分布，在“生产可能性边界（production possibility frontier）”框架下可视化了性状间的权衡关系。本研究结果表明，部分功能丧失型突变可提升体外培养的菌体生长速率或发光强度，但单一性状的提升通常会以另一性状的削弱为代价。