Regulation of l -Lactate Utilization by the FadR-Type Regulator LldR of Corynebacterium glutamicum

Abstract

Corynebacterium glutamicum can grow on L-lactate as a sole carbon and energy source. The NCgl2816-lldD operon encoding a putative transporter (NCgl2816) and a quinone-dependent L-lactate dehydrogenase (LldD) is required for L-lactate utilization. DNA affinity chromatography revealed that the FadR-type regulator LldR (encoded by NCgl2814) binds to the upstream region of NCgl2816-lldD. Overexpression of lldR resulted in strongly reduced NCgl2816-lldD mRNA levels and strongly reduced LldD activity, and as a consequence, a severe growth defect was observed in cells grown on L-lactate as the sole carbon and energy source, but not in cells grown on glucose, ribose, or acetate. Deletion of lldR had no effect on growth on these carbon sources but resulted in high NCgl2816-lldD mRNA levels and high LldD activity in the presence and absence of L-lactate. Lactate is a major product of anaerobic metabolism, but it also serves as a carbon and energy source for anaerobic and aerobic microorganisms. Lactate can be fermented to acetate, propionate, or butyrate by, e.g., sulfate-reducing bacteria, propionibacteria, or Eubacterium hallii (11). When oxygen becomes available but glucose is limiting, Lactobacillus plantarum converts its fermentation product, lactate, to acetate (18). Aerobic growth with L-lactate as the sole carbon and energy source has been studied in Escherichia coli in some detail. L-Lactate is taken up into the E. coli cell either by the L-lactate permease LldP or by the glycolate permease GlcA (39). LLactate is oxidized to the central metabolite pyruvate by quinone-dependent L-lactate dehydrogenase (LldD; EC 1.1.2.3) (10). For growth on L-lactate, E. coli requires lldD, which forms an operon with lldP and the putative lactate regulator gene lldR (10). Transcription of lldDRP is repressed by ArcAB under anaerobic reducing conditions (24) and is maximal in the presence of L-lactate (10); however, regulation of lldDRP by the putative regulator LldR encoded in this operon has not been analyzed yet in detail.Recently, we identified the L-lactate utilization operon in Corynebacterium glutamicum, a nonpathogenic gram-positive soil bacterium that is widely used for biotechnological production of amino acids such as L-glutamate and L-lysine. C. glutamicum can grow aerobically on a variety of sugars, sugar alcohols, and organic acids, including L-lactate, as sole carbon and energy sources (9,17,27,31,36,59). C. glutamicum forms L-lactate with the soluble NAD ϩ -dependent L-lactate dehydrogenase (EC 1.1.1.27) encoded by ldhA (3, 22) under oxygen deprivation (22) and as a by-product during glutamate and lysine production (27,28,53). For L-lactate utilization, on the other hand, C. glutamicum requires the quinone-dependent L-lactate dehydrogenase LldD (EC 1.1.2.3) (53), which is a peripheral membrane protein (51) catalyzing oxidation of Llactate to pyruvate (3, 53).The C. glutamicum L-lactate utilization operon comprises the quinone-dependent L-lactate dehydrogenase gene lldD and a gene encoding a putative permease (NCgl281...

A role of the transcriptional regulator LldR (NCgl2814) in glutamate metabolism under biotin-limited conditions in Corynebacterium glutamicum

“…4a). It was also reported that overexpression and deletion of lldR have no significant effects on growth rate or biomass formation compared to WT in minimal medium containing glucose as a sole carbon source (Georgi et al, 2008). In the lldR disruptant strain, glutamate production was slightly increased reproducibly, although it was not significant statistically.…”

“…Moreover, LldR also represses the fruR-fruk-ptsF operon responsible for fructose utilization (Gao et al, 2008). It was reported that overexpression of lldR resulted in a decrease in expression of ldhA and the NCgl2816-lldD operon, while disruption of lldR showed a significant increase in expression of the NCgl2816-lldD operon (Gao et al, 2008;Georgi et al, 2008). Disruption of lldR showed a significant increase in ldhA expression in the absence of sugar (Gao et al, 2008) and showed no significant change in ldhA expression in the presence of sugar (Georgi et al, 2008).…”

“…lldR (NCgl2814) encodes a GntR-type transcriptional repressor of the NCgl2816-lldD operon. L-Lactate binds to LldR, preventing repression of NCgl2816-lldD by LldR (Gao et al, 2008;Georgi et al, 2008). LldR also acts as a repressor of ldhA in the absence of L-lactate, but ldhA expression is primarily repressed by the DeoR-type transcription regulator, SugR, in the absence of sugar Toyoda et al, 2009a, b).…”

Rv0494 is a starvation-inducible, auto-regulatory FadR-like regulator from Mycobacterium tuberculosis

“…FadR regulators are the most abundant amongst the GntR family of transcription regulators and consist of all a-helical C-terminal domains with a mean length of~160 aa (Haydon & Guest, 1991;Rigali et al, 2002;Vindal et al, 2007a, b). Extensive studies have been carried on the FadRs of Escherichia coli, Vibrio vulnificus and Corynebacterium glutamicum, and they are reported to play important roles both in cellular physiology and virulence (Agari et al, 2011;Brown & Gulig, 2008;Casali et al, 2006;Georgi et al, 2008;Santangelo et al, 2008). However, current knowledge on the promoter architecture and regulation of these FadR-like proteins in mycobacteria is limited.…”

Transcriptional regulation of the l-lactate permease gene lutP by the LutR repressor of Bacillus subtilis RO-NN-1

“…An imperfect inverted repeat sequence 59-TNGT-N 3 -ACNA-39 can serve as an operator for the LldR repressor (Gao et al, 2008;Georgi et al, 2008). The lutABC operon of the undomesticated strain NCIB 3610 of B. subtilis encodes three iron-sulfur-containing proteins responsible for L-lactate utilization (Chai et al, 2009).…”