(176ag) Exploring ATP Dynamics for Metabolic Engineering

Adenosine-5’-triphosphate (ATP), the primary energy currency in cellular processes, plays a pivotal role in driving metabolic activities and the production of various molecules. Despite its importance, the influence of intracellular ATP dynamics on bioproduction and methods to exploit ATP dynamics for enhanced bioproduction remain largely unexplored.

Here, we harnessed an ATP biosensor to dissect ATP dynamics across different growth phases and varying carbon sources in multiple microbial strains. We found transient ATP accumulations during the transition from exponential to early stationary growth phases in multiple conditions. The observed ATP dynamics coincide with fatty acid (FA) production in Escherichia coli and polyhydroxyalkanoate (PHA) production in Pseudomonas putida. Importantly, we discovered unexpectedly higher ATP levels in E. coli cells grown in acetate, which is traditionally considered an unfavorable carbon source for E. coli fermentation. The addition of this cheap and widely available carbon source substantially enhanced fatty acid bioproduction. We further demonstrated that similar strategies based on ATP dynamics boosted PHA production in engineered P. putida strains. Moreover, we employed ATP dynamics monitoring as a sensitive diagnostic tool to assess metabolic burden within engineered microbial cells, revealing bottleneck steps that limit limonene bioproduction.

This work not only elucidates the fundamental relationship between ATP dynamics and microbial bioproduction but also showcases its value in enhancing the production of a wide range of chemicals, proteins, and materials in various microbial species. Our results challenge conventional assumptions by revealing higher ATP concentrations in E. coli cells grown in acetate than in glucose. We provided a simple and cost-effective approach to enhance bioproduction through supplementation of ATP-promoting carbon sources. Our study also has broad implications for understanding microbial energy homeostasis, optimizing bioproduction processes, and identifying sources of metabolic burden. These insights can contribute to diverse fields, including microbiology, metabolic engineering, biotechnology, and chemical biology.