An acetoin-regulated expression system of Bacillussubtilis

Abstract

An expression system, which is based on the promoter of the acoABCL operon of Bacillus subtilis was developed and characterized. The acoABCL operon codes for the acetoin dehydrogenase complex, which is the major enzyme system responsible for the catabolism of acetoin in B. subtilis. Besides weak organic acids, the neutral overflow metabolite acetoin is metabolized by the cells in the early stationary phase. Transcription of reporter gene fusions with the acoA promoter of this operon is strongly repressed by glucose but induced by acetoin as soon as the preferred carbon source glucose is exhausted. The co-expression of an additional copy of the regulator gene acoR led to more than twofold higher activity of the acoA promoter. It is demonstrated that the induction of this promoter in growing cells with acetoin is possible with non-phosphotransferase system sugars as carbon and energy source and in a ccpA mutant background. Moreover, it could be shown that the activity of the acoA-directed expression system correlates with the level of acetoin in the medium. During glucose limitation, the utilization of the alternative energy source acetoin keeps the protein synthesis machinery of B. subtilis cells active and thus allows for a long lasting acoA-controlled expression of recombinant genes.

Bet-hedging and epigenetic inheritance in bacterial cell development

“…1B). This secondary growth phase is probably caused by the utilization of overflow metabolites, such as acetoin, after the disappearance of glucose (13). After the cessation of the secondary growth phase, there is a period of Ϸ10 h during which there is no cell growth.…”

“…Because spore formation is a long and energy-intensive process, it is plausible that sporulating cells use the overflow metabolites to complete this cell fate. The induction of overflow metabolism can be monitored by following the expression of the acetoin catabolic pathway (acoA-L), which is exclusively activated when the growth medium is exhausted for glucose (13). Indeed, we found that all cells in a microcolony express this operon at the end of the logarithmic growth phase (SI Fig.…”

The Bacillus subtilis GntR Family Repressor YtrA Responds to Cell Wall Antibiotics

“…This assignment was based on a slight decrease in the rate of acetoin reutilization in a ytrA operon deletion. However, the ytrA operon was not induced by acetoin (45), and subsequent studies indicate that acetoin catabolism is determined by the carbon metabolite-repressed and acetoin-inducible acoABCL operon (1,35,41). Conversely, induction of the ytrA operon was noted in previous global analyses of antibiotic stress responses, and ytrA was even proposed as a reporter for glycopeptide antibiotics (18).…”

“…It is unknown whether YtrBCDEF acts as one integrated or two separate ABC transport systems (32,45). Although initially suggested to encode an uptake system for acetoin, based on a modest decrease in acetoin utilization in a mutant strain (45), more recent studies indicate that acetoin catabolism is determined by the acetoin-inducible acoABCL operon (35,41). Moreover, acetoin does not induce ytrA expression (45).The ywoBCD operon encodes a protein of unknown function, a cysteine hydrolase, and an MFS family major facilitator transporter, respectively.…”

Bioscouring and bleaching process of cotton fabrics – an opportunity of saving water and energy

“…Screening approaches are being performed to rapidly identify enzymes with potential industrial application (Korf et al, 2005). For this purpose, different expression hosts (Esherichia coli, Bacillus sp., Sacharomyces cerevisae, Pichia pastoris) have been developed to express heterologous proteins (Chelikani, Reeves, Rajbhandary, & Khorana, 2006;Silbersack et al, 2006;Li et al, 2007).…”