Integrated Regulation of Acetoin Fermentation by Quorum Sensing and pH in Serratia plymuthica RVH1

Abstract

During fermentation of sugars, a number of bacterial species are able to switch from mixed acid production to acetoin and 2,3-butanediol production in order to avoid lethal acidification of their environment, although the regulation of this switch is only poorly understood. In this study, we report the identification of the budAB structural operon, involved in acetoin production in Serratia plymuthica RVH1, and its activation by a LysRtype regulator encoded by budR, immediately upstream of this operon. In addition, the regulation of budR transcription was elucidated and found to be subject to negative control by BudR itself and to positive control by external stimuli such as N-(3-oxohexanoyl)-L-homoserine lactone (OHHL) quorum sensing signaling molecules and acetate. Interestingly, however, we observed that induction of budR transcription by OHHL or acetate did not require BudR, indicating the involvement of additional regulatory factors in relaying these environmental signals to the budR promoter.A wide variety of strains are able to switch to acetoin and 2,3-butanediol production during fermentative growth to counteract the lethal acidification that typically accompanies mixed acid production (3,17,21). Butanediol is produced by first converting two molecules of pyruvate into ␣-acetolactate by ␣-acetolactate synthase, after which ␣-acetolactate is processed to acetoin by ␣-acetolactate decarboxylase. Finally, acetoin can be converted reversibly to 2,3-butanediol through the action of acetoin reductase (25). Often, the structural genes for 2,3-butanediol production are organized into an operon that is controlled by a neighboring and divergently transcribed LysR homologue (6,12,15). Nevertheless, the regulatory aspects, and especially the integration of environmental cues into the control of this pathway, remain largely unexplored.In this article, we report on the identification and regulation of the acetoin operon in Serratia plymuthica RVH1 and on its activation at the molecular level by signals of acidification and quorum sensing (QS). MATERIALS AND METHODSStrains and growth conditions. Bacterial strains and plasmids used in this study are listed in Table 1. Escherichia coli and S. plymuthica RVH1 strains were cultured at 30°C in Luria-Bertani (LB) broth or agar (1.5% agar) or in M9 minimal medium (14). Where necessary, LB medium was acidified using 1 M HCl or buffered with 100 mM phosphate buffer at pH 7.0. As described previously (24), growth was monitored using a Bioscreen C microbiology reader (Thermo Life Sciences, Brussels, Belgium), while acetoin production and medium acidification were determined by the Voges-Proskauer and methyl red assays, respectively.The following concentrations of sugar and/or antibiotics (Applichem, Darmstadt, Germany) were used when appropriate: glucose, 0.5%; ampicillin, 100 g/ml; carbenicillin, 300 g/ml; kanamycin, 50 g/ml; and gentamicin, 20 g/ml. The synthetic N-acyl-L-homoserine lactone (AHL) used in this study, N-(3-oxohexanoyl)-L-homoserine lactone (OHHL) (5 M solution i...

HPLC and UPLC Analyses of Acetoin in Bacterial Culture Fluid

“…It is one of thirty basic chemical components that the U.S. Department of Energy has prioritized for development and utilization [3]. [4][5][6].…”

(p)ppGpp, a Small Nucleotide Regulator, Directs the Metabolic Fate of Glucose in Vibrio cholerae

“…This finding suggested that the QS system likely plays a role in controlling glucose metabolism in V. cholerae. Similarly, the level of acetoin fermentation was also regulated by QS in Serratia plymuthica (19). Detailed signaling cues and molecular determinants leading to the activation of acetoin fermentation, however, are not fully understood.…”

Acetoin Synthesis Acquisition Favors Escherichia coli Growth at Low pH

“…In fact, the latter bacteria follow the mixed-acid fermentation route during the early stages of growth and switch to the 2,3-butanediol route in the late-exponential phase (3,4). This switch is generally viewed as a strategy of these bacteria to prevent excessive medium acidification, and it has been demonstrated that knockout of the 2,3-butanediol pathway indeed results in more-pronounced acidification, early growth arrest, and cell death (5,6).…”