Horizontal transfer of a pathway for coumarate catabolism unexpectedly inhibits purine nucleotide biosynthesis

Abstract

Metabolic pathways are frequently transferred between bacterial strains in the environment through horizontal gene transfer (HGT), yet laboratory engineering to introduce new metabolic pathways often fails. Successful use of a pathway requires co-evolution of both pathway and host, and these interactions may be disrupted upon transfer to a new host. Here we show that two different pathways for catabolism of coumarate failed to function when initially transferred into Escherichia coli. Using laboratory evolution, we elucidated the factors limiting activity of the newly-acquired pathways and the modifications required to overcome these limitations. Both pathways required mutations to the host to enable effective growth with coumarate, but the necessary mutations differed depending on the chemistry and intermediates of the pathways. In one case, an intermediate inhibited purine nucleotide biosynthesis, and this inhibition was relieved by single amino acid mutations to IMP dehydrogenase. A strain that natively contains this coumarate catabolism pathway, Acinetobacter baumannii, is already resistant to inhibition by the relevant intermediate, suggesting that natural pathway transfers have faced and overcome similar challenges. These discoveries will aid in our understanding of HGT and ability to predictably engineer metabolism.

Modular Engineering of Biomass Degradation Pathways

“…For example, the implantation of two coumarate catabolic pathways into E. coli failed until laboratory evolution revealed a unique mechanism of interference. In one pathway, an intermediate was found to inhibit purine nucleotide biosynthesis [27].…”

“…However, the introduction of new metabolic pathways, such as those for catabolism of lignocellulosic sugars and aromatic compounds, can be a significant perturbation to the metabolic and regulatory networks of the host. For example, heterologous pathways may produce new metabolites that interact with native enzymes and transcription factors, or heterologous enzymes can divert native metabolites into nonproductive pathways [26,27]. Understanding the challenges involved in heterologous pathway expression will allow us to engineer new metabolic modules that minimize these impacts [28].…”