Identification and regulation of an alternative PTS for disaccharide utilization in Clostridium acetobutylicum.

Abstract

Clostridium acetobutylicum is an important solventogenic bacterium capable of acetone-butanol-ethanol fermentation by utilizing a variety of carbon sources. It employs the transport systems of the phosphoenolpyruvate:sugar phosphotransferase systems (PTSs) to assimilate various saccharides. Here, we investigated a β-glucoside PTS (bglT) encoded by the bgl operon (bglGTH) in C. acetobutylicum, which showed significant expression in response to cellobiose and sucrose. Interestingly, bglT is not essential for the transport of these sugars, as C. acetobutylicum possesses dedicated PTSs for the uptake of each individual sugar. We further elucidated the regulatory mechanism of bglT, which is governed by an upstream anti-transcriptional termination factor (bglG). A putative ribonucleic antiterminator (RAT) was identified upstream of bglG and bglT. Inactivation of bglG led to consistent read-through frequencies of the genes downstream of the RAT, irrespective of the sugar present. Conversely, complete removal of RAT elevated the transcriptional levels of downstream genes, while partial deletion of RAT, causing a long stem-loop structure (terminator), resulted in transcription termination. These findings provide novel insights into the regulatory mechanisms controlling sugar utilization in C. acetobutylicum.

Importance: Cellulose, the most abundant organic compound on Earth, is primarily found in plant cell walls and can be broken down into sugars such as cellobiose. These sugars are crucial for microbial fermentation, especially in biofuel production. Clostridium acetobutylicum, a promising microorganism for producing short-chain alcohol chemicals, can utilize cellulose degradation products as a carbon source for fermentation. This study identifies the transport systems involved in the utilization of cellobiose and other disaccharides in C. acetobutylicum and analyzes their regulatory mechanisms. Understanding these pathways is essential for enhancing biofuel production from plant biomass.