An Exporter of Leucine, LeuE Functions as an Exporter of 1-Aminocyclopropane-1-Carboxylic Acid in Escherichia coli

1-aminocyclopropane-1-carboxylic acid (ACC), a crucial intermediate in the ethylene biosynthetic pathway, plays a decisive role in regulating plant developmental processes and stress responses. In heterologous biosynthesis systems, ACC biosynthesis is inhibited due to its antibacterial activity. Enhancing ACC export confers advantages to ACC-producing biotechnological platforms and microbial cell factories. While bacterial transporters for canonical amino acids in Corynebacterium glutamicum and Escherichia coli have been extensively studied, their roles in ACC transport remain unexplored. To identify the exporters mediating ACC transport in E. coli , relevant experiments were performed in this study. The results showed that the Leucine-exporter LeuE acts as a high-efficiency ACC exporter in E. coli . Subsequently, structural modeling coupled with site-directed mutagenesis revealed that L160 is a critical residue in determining the ACC transport specificity of LeuE. The L160Y mutation enhanced E. coli tolerance to ACC, elevating the threshold from 50   mM to 100   mM. Structural modeling predicted that the L160Y mutation-induced compaction reduced the transporter channel diameter by 28% (from 9.551   Å to 6.876   Å), a change that may enhance the binding affinity of ACC. Notably, a significant reduction in intracellular ACC levels was observed in both the LeuE-overexpressing and LeuE (L160Y)-overexpressing strains. These findings enable large-scale biosynthesis of ACC and its derivatives via engineered microbial systems, thereby facilitating cost-effective development of ACC-based pharmaceuticals and agrochemicals.