Engineering genetic regulation in Clostridium to improve industrial properties

Abstract

Clostridium are anaerobic Gram-positive bacteria with members innately able to produce a variety of commercially valuable chemicals via fermentation from renewable or waste feedstocks. Although this method is more sustainable and carbon-neutral than production from petrochemical sources, improvements are required to make fermentation financially competitive. Here, a novel strain engineering technique, gene- network rewiring, is used to improve industrial properties of Clostridium acetobutylicum. This approach is non-laborious, well tolerated and requires minimal knowledge of the systems governing the phenotypes. The transcription factor network consists of nodes (transcription factors) and edges (gene regulation). Endogenous transcription factor genes are separated between their coding sequences and promoters, and then randomly combinatorially assembled, generating new nodes within the network. This generates a novel combination of output signal (gene expression) from input signal (promoter regulation), thereby changing the way that the organism senses and responds to stimuli. Here, rewiring events have been selected that improve tolerance to the lignocellulose feedstock inhibitors furfural and hydroxymethylfurfural (HMF).