Characterisation of a glucose phosphotransferase system in Clostridium acetobutylicum ATCC 824

Abstract

The transport of glucose by the solventogenic anaerobe Clostridium acetobutylicum was investigated. Glucose phosphoenolpyruvate (PEP)-dependent phosphotransferase system (PTS) activity was detected in extracts prepared from cultures grown on glucose and extract fractionation revealed that both soluble and membrane components are required for activity. Glucose PTS activity was inhibited by the analogue methyl alpha-glucoside, indicating that the PTS enzyme II belongs to the glucose-glucoside (Glc) family of proteins. Consistent with this conclusion, labelled methyl alpha-glucoside was phosphorylated by PEP in cell-free extracts and this activity was inhibited by glucose. A single gene encoding a putative enzyme II of the glucose family, which we have designated glcG, was identified from the C. acetobutylicum ATCC 824 genome sequence. In common with certain other low-GC gram-positive bacteria, including Bacillus subtilis, the C. acetobutylicum glcG gene appears to be associated with a BglG-type regulator mechanism, as it is preceded by a transcription terminator that is partially overlapped by a typical ribonucleic antiterminator (RAT) sequence, and is downstream of an open reading frame that appears to encode a transcription antiterminator protein. This is the first report of a glucose transport mechanism in this industrially important organism.

Confirmation and Elimination of Xylose Metabolism Bottlenecks in Glucose Phosphoenolpyruvate-Dependent Phosphotransferase System-Deficient Clostridium acetobutylicum for Simultaneous Utilization of Glucose, Xylose, and Arabinose

“…PEPdependent glucose PTS activity has been detected in C. acetobutylicum ATCC 824 (45), indicating that this activity may play a leading role for D-glucose metabolism in this anaerobe. To attenuate glucose PTS activity in cells, gene glcG (cac0570), which was suggested by bioinformatics analysis to encode the PTS enzyme II (45), was disrupted using TargeTron technology (42).…”

“…In the present study, a new strategy that enabled C. acetobutylicum to efficiently coferment D-glucose, D-xylose, and L-arabinose is presented. We first inactivated the glcG gene (45), which greatly reduced "glucose repression" of D-xylose and L-arabinose metabolism. Next, the rate-limiting steps of the D-xylose pathway were confirmed, and the bottleneck was relieved.…”

Combination of RNA sequencing and metabolite data to elucidate improved toxic compound tolerance and butanol fermentation of Clostridium acetobutylicum from wheat straw hydrolysate by supplying sodium sulfide

“…It was induced by the addition of SS, and in the presence of SS, its expression level was 8.26 times greater than in the unsupplemented control. A second putative glucose PTS is encoded by two divergently transcribed genes, CAC1353 and CAC1354, which encode PTS IICB and PTS IIA components, respectively (Tangney and Mitchell, 2007). Nevertheless, neither of these genes displayed significantly altered expression in the presence of SS (Fig.…”

Cellodextrin and Laminaribiose ABC Transporters in Clostridium thermocellum

“…Sugar transport in microorganisms is usually mediated by phosphoenolpyruvate-dependent phosphotransferase systems, ATP-binding cassette (ABC) transport systems, or proton-linked transport systems. In Clostridium acetobutylicum, the transport of glucose and lactose is mediated by the phosphotransferase system (33,39), whereas in Streptomyces reticuli, another gram-positive soil bacterium, the transport of cellobiose and cellotriose is mediated by ABC transporters (26). Strobel et al have suggested that in C. thermocellum, cellodextrins enter the cells via ATP-dependent transport systems, i.e., the ABC transporters (31).…”