scholarly journals The ldhA Gene, Encoding Fermentative l-Lactate Dehydrogenase of Corynebacterium glutamicum, Is under the Control of Positive Feedback Regulation Mediated by LldR

2009 ◽  
Vol 191 (13) ◽  
pp. 4251-4258 ◽  
Author(s):  
Koichi Toyoda ◽  
Haruhiko Teramoto ◽  
Masayuki Inui ◽  
Hideaki Yukawa
Keyword(s):  
Lactate Dehydrogenase ◽  
Corynebacterium Glutamicum ◽  
Promoter Region ◽  
Lactate Production ◽  
Feedback Regulation ◽  
Gene Encoding ◽  
Key Enzyme ◽  
In The Wild ◽  
Lactate Utilization ◽  
Ldha Gene

ABSTRACT Corynebacterium glutamicum ldhA encodes l-lactate dehydrogenase, a key enzyme that couples l-lactate production to reoxidation of NADH formed during glycolysis. We previously showed that in the absence of sugar, SugR binds to the ldhA promoter region, thereby repressing ldhA expression. In this study we show that LldR is another protein that binds to the ldhA promoter region, thus regulating ldhA expression. LldR has hitherto been characterized as an l-lactate-responsive transcriptional repressor of l-lactate utilization genes. Transposon mutagenesis of a reporter strain carrying a chromosomal ldhA promoter-lacZ fusion (PldhA-lacZ) revealed that ldhA disruption drastically decreased expression of PldhA-lacZ. PldhA-lacZ expression in the ldhA mutant was restored by deletion of lldR, suggesting that LldR acts as a repressor of ldhA in the absence of l-lactate and the LldR-mediated repression is not relieved in the ldhA mutant due to its inability to produce l-lactate. lldR deletion did not affect PldhA-lacZ expression in the wild-type background during growth on either glucose, acetate, or l-lactate. However, it upregulated PldhA-lacZ expression in the sugR mutant background during growth on acetate. The binding sites of LldR and SugR are located around the −35 and −10 regions of the ldhA promoter, respectively. C. glutamicum ldhA expression is therefore primarily repressed by SugR in the absence of sugar. In the presence of sugar, SugR-mediated repression of ldhA is alleviated, and ldhA expression is additionally enhanced by LldR inactivation in response to l-lactate produced by LdhA.

scholarly journals The ldhA Gene Encoding Fermentative L-Lactate Dehydrogenase in Corynebacterium glutamicum Is Positively Regulated by the Global Regulator GlxR

2021 ◽  
Vol 9 (3) ◽  
pp. 550
Author(s):  
Koichi Toyoda ◽  
Masayuki Inui
Keyword(s):  
Lactate Dehydrogenase ◽  
Binding Site ◽  
Corynebacterium Glutamicum ◽  
Lactate Production ◽  
Exponential Phase ◽  
Aerobic Bacterium ◽  
Global Regulator ◽  
Gene Encoding ◽  
Fermentation Products ◽  
Ldha Gene

Bacterial metabolism shifts from aerobic respiration to fermentation at the transition from exponential to stationary growth phases in response to limited oxygen availability. Corynebacterium glutamicum, a Gram-positive, facultative aerobic bacterium used for industrial amino acid production, excretes L-lactate, acetate, and succinate as fermentation products. The ldhA gene encoding L-lactate dehydrogenase is solely responsible for L-lactate production. Its expression is repressed at the exponential phase and prominently induced at the transition phase. ldhA is transcriptionally repressed by the sugar-phosphate-responsive regulator SugR and L-lactate-responsive regulator LldR. Although ldhA expression is derepressed even at the exponential phase in the sugR and lldR double deletion mutant, a further increase in its expression is still observed at the stationary phase, implicating the action of additional transcription regulators. In this study, involvement of the cAMP receptor protein-type global regulator GlxR in the regulation of ldhA expression was investigated. The GlxR-binding site found in the ldhA promoter was modified to inhibit or enhance binding of GlxR. The ldhA promoter activity and expression of ldhA were altered in proportion to the binding affinity of GlxR. Similarly, L-lactate production was also affected by the binding site modification. Thus, GlxR was demonstrated to act as a transcriptional activator of ldhA.

scholarly journals Characterization of a Corynebacterium glutamicum Lactate Utilization Operon Induced during Temperature-Triggered Glutamate Production

2005 ◽  
Vol 71 (10) ◽  
pp. 5920-5928 ◽  
Author(s):  
Corinna Stansen ◽  
Davin Uy ◽  
Stephane Delaunay ◽  
Lothar Eggeling ◽  
Jean-Louis Goergen ◽  
...  
Keyword(s):  
Lactate Dehydrogenase ◽  
Carbon Source ◽  
Corynebacterium Glutamicum ◽  
Specific Activity ◽  
Temperature Shift ◽  
Pcr Analysis ◽  
Mrna Levels ◽  
Reverse Transcription Pcr ◽  
Glutamate Production ◽  
Lactate Utilization

ABSTRACT Gene expression changes of glutamate-producing Corynebacterium glutamicum were identified in transcriptome comparisons by DNA microarray analysis. During glutamate production induced by a temperature shift, C. glutamicum strain 2262 showed significantly higher mRNA levels of the NCgl2816 and NCgl2817 genes than its non-glutamate-producing derivative 2262NP. Reverse transcription-PCR analysis showed that the two genes together constitute an operon. NCgl2816 putatively codes for a lactate permease, while NCgl2817 was demonstrated to encode quinone-dependent l-lactate dehydrogenase, which was named LldD. C. glutamicum LldD displayed Michaelis-Menten kinetics for the substrate l-lactate with a Km of about 0.51 mM. The specific activity of LldD was about 10-fold higher during growth on l-lactate or on an l-lactate-glucose mixture than during growth on glucose, d-lactate, or pyruvate, while the specific activity of quinone-dependent d-lactate dehydrogenase differed little with the carbon source. RNA levels of NCgl2816 and lldD were about 18-fold higher during growth on l-lactate than on pyruvate. Disruption of the NCgl2816-lldD operon resulted in loss of the ability to utilize l-lactate as the sole carbon source. Expression of lldD restored l-lactate utilization, indicating that the function of the permease gene NCgl2816 is dispensable, while LldD is essential, for growth of C. glutamicum on l-lactate.

scholarly journals Lactate Dehydrogenase Is the Key Enzyme for Pneumococcal Pyruvate Metabolism and Pneumococcal Survival in Blood

2014 ◽  
Vol 82 (12) ◽  
pp. 5099-5109 ◽  
Author(s):  
Paula Gaspar ◽  
Firas A. Y. Al-Bayati ◽  
Peter W. Andrew ◽  
Ana Rute Neves ◽  
Hasan Yesilkaya
Keyword(s):  
Lactate Dehydrogenase ◽  
Lactate Production ◽  
Virulence Gene ◽  
Redox Balance ◽  
Central Metabolism ◽  
Content Type ◽  
Virulence Gene Expression ◽  
Key Enzyme

ABSTRACTStreptococcus pneumoniaeis a fermentative microorganism and causes serious diseases in humans, including otitis media, bacteremia, meningitis, and pneumonia. However, the mechanisms enabling pneumococcal survival in the host and causing disease in different tissues are incompletely understood. The available evidence indicates a strong link between the central metabolism and pneumococcal virulence. To further our knowledge on pneumococcal virulence, we investigated the role of lactate dehydrogenase (LDH), which converts pyruvate to lactate and is an essential enzyme for redox balance, in the pneumococcal central metabolism and virulence using an isogenicldhmutant. Loss of LDH led to a dramatic reduction of the growth rate, pinpointing the key role of this enzyme in fermentative metabolism. The pattern of end products was altered, and lactate production was totally blocked. The fermentation profile was confirmed byin vivonuclear magnetic resonance (NMR) measurements of glucose metabolism in nongrowing cell suspensions of theldhmutant. In this strain, a bottleneck in the fermentative steps is evident from the accumulation of pyruvate, revealing LDH as the most efficient enzyme in pyruvate conversion. An increase in ethanol production was also observed, indicating that in the absence of LDH the redox balance is maintained through alcohol dehydrogenase activity. We also found that the absence of LDH renders the pneumococci avirulent after intravenous infection and leads to a significant reduction in virulence in a model of pneumonia that develops after intranasal infection, likely due to a decrease in energy generation and virulence gene expression.

scholarly journals Elucidating the Role and Regulation of a Lactate Permease as Lactate Transporter in Bacillus coagulans DSM1

2019 ◽  
Vol 85 (14) ◽  
Author(s):  
Yu Wang ◽  
Caili Zhang ◽  
Guoxia Liu ◽  
Jiansong Ju ◽  
Bo Yu ◽  
...  
Keyword(s):  
Intergenic Region ◽  
Lactate Production ◽  
Bacillus Coagulans ◽  
Upstream Sequence ◽  
Gene Encoding ◽  
Content Type ◽  
Lactate Uptake ◽  
Lactate Utilization ◽  
Lactate Transporter ◽  
Encoding Gene

ABSTRACT A key feature of Bacillus coagulans is its ability to produce l-lactate via homofermentative metabolism. A putative lactate permease-encoding gene (lutP) and the gene encoding its regulator (lutR) were identified in one operon in B. coagulans strains. LutP orthologs are highly conserved and located adjacent to the gene cluster related to lactate utilization in most lactate-utilizing microorganisms. However, no lactate utilization genes were found adjacent to lutP in all sequenced B. coagulans strains. The stand-alone presence of lutP in l-lactate producers indicates that it may have functions in lactate production. In this study, B. coagulans DSM1 was used as a representative strain, and the critical roles of LutP and its regulation were described. Transport property assays showed that LutP was essential for lactate uptake. Its regulator LutR directly interacted with the lutP-lutR intergenic region, and lutP transcription was activated by l-lactate via regulation by LutR. A biolayer interferometry assay further confirmed that LutR bound to an 11-bp inverted repeat in the intergenic region, and lutP transcription began when the binding of LutR to the lutP upstream sequence was inhibited. We conclusively showed that lutP encodes a functional lactate permease in B. coagulans. IMPORTANCE Lactate-utilizing strains require lactate permease (LutP) to transport lactate into cells. Bacillus coagulans LutP is a previously uncharacterized lactate permease with no lactate utilization genes situated either adjacent to or remotely from it. In this study, an active lactate permease in an l-lactate producer, B. coagulans DSM1, was identified. Lactate supplementation regulated the expression of lactate permease. This study presents physiological evidence of the presence of a lactate transporter in B. coagulans. Our findings indicate a potential target for the engineering of strains in order to improve their fermentation characteristics.

scholarly journals The Global Repressor SugR Controls Expression of Genes of Glycolysis and of the l-Lactate Dehydrogenase LdhA in Corynebacterium glutamicum

2008 ◽  
Vol 190 (24) ◽  
pp. 8033-8044 ◽  
Author(s):  
Verena Engels ◽  
Steffen N. Lindner ◽  
Volker F. Wendisch
Keyword(s):  
Lactate Dehydrogenase ◽  
Corynebacterium Glutamicum ◽  
Target Genes ◽  
Lactate Production ◽  
Oxygen Deprivation ◽  
Mrna Levels ◽  
Phosphotransferase System ◽  
Gel Retardation ◽  
Activity Measurements

ABSTRACT The transcriptional regulator SugR from Corynebacterium glutamicum represses genes of the phosphoenolpyruvate-dependent phosphotransferase system (PTS). Growth experiments revealed that the overexpression of sugR not only perturbed the growth of C. glutamicum on the PTS sugars glucose, fructose, and sucrose but also led to a significant growth inhibition on ribose, which is not taken up via the PTS. Chromatin immunoprecipitation combined with DNA microarray analysis and gel retardation experiments were performed to identify further target genes of SugR. Gel retardation analysis confirmed that SugR bound to the promoter regions of genes of the glycolytic enzymes 6-phosphofructokinase (pfkA), fructose-1,6-bisphosphate aldolase (fba), enolase (eno), pyruvate kinase (pyk), and NAD-dependent l-lactate dehydrogenase (ldhA). The deletion of sugR resulted in increased mRNA levels of eno, pyk, and ldhA in acetate medium. Enzyme activity measurements revealed that SugR-mediated repression affects the activities of PfkA, Fba, and LdhA in vivo. As the deletion of sugR led to increased LdhA activity under aerobic and under oxygen deprivation conditions, l-lactate production by C. glutamicum was determined. The overexpression of sugR reduced l-lactate production by about 25%, and sugR deletion increased l-lactate formation under oxygen deprivation conditions by threefold. Thus, SugR functions as a global repressor of genes of the PTS, glycolysis, and fermentative l-lactate dehydrogenase in C. glutamicum.

scholarly journals Regulation of Alginate Biosynthesis inPseudomonas syringae pv. syringae

1999 ◽  
Vol 181 (11) ◽  
pp. 3478-3485 ◽  
Author(s):  
Mohamed K. Fakhr ◽  
Alejandro Peñaloza-Vázquez ◽  
Ananda M. Chakrabarty ◽  
Carol L. Bender
Keyword(s):  
Sigma Factor ◽  
Promoter Region ◽  
Biosynthetic Pathway ◽  
Response Regulator ◽  
Consensus Sequence ◽  
Gene Encoding ◽  
Environmental Signals ◽  
Alginate Production ◽  
Key Enzyme ◽  
Alginate Biosynthesis

ABSTRACT Both Pseudomonas aeruginosa and the phytopathogenP. syringae produce the exopolysaccharide alginate. However, the environmental signals that trigger alginate gene expression in P. syringae are different from those inP. aeruginosa with copper being a major signal in P. syringae. In P. aeruginosa, the alternate sigma factor encoded by algT (ς22) and the response regulator AlgR1 are required for transcription of algD, a gene which encodes a key enzyme in the alginate biosynthetic pathway. In the present study, we cloned and characterized the gene encoding AlgR1 from P. syringae. The deduced amino acid sequence of AlgR1 from P. syringae showed 86% identity to its P. aeruginosa counterpart. Sequence analysis of the region flankingalgR1 in P. syringae revealed the presence ofargH, algZ, and hemC in an arrangement virtually identical to that reported in P. aeruginosa. An algR1 mutant, P. syringaeFF5.32, was defective in alginate production but could be complemented when algR1 was expressed in trans. ThealgD promoter region in P. syringae(PsalgD) was also characterized and shown to diverge significantly from the algD promoter in P. aeruginosa. Unlike P. aeruginosa, algR1was not required for the transcription of algD in P. syringae, and PsalgD lacked the consensus sequence recognized by AlgR1. However, both the algD andalgR1 upstream regions in P. syringae contained the consensus sequence recognized by ς22, suggesting thatalgT is required for transcription of both genes.

Regulation of the ldhA gene, encoding the fermentative lactate dehydrogenase of Escherichia coli

Microbiology ◽  
2001 ◽  
Vol 147 (9) ◽  
pp. 2437-2446 ◽  
Author(s):  
Gene Ruijun Jiang ◽  
Sonia Nikolova ◽  
David P Clark
Keyword(s):  
Escherichia Coli ◽  
Lactate Dehydrogenase ◽  
Gene Encoding ◽  
Ldha Gene

scholarly journals Expression and function of the cdgD gene, encoding a CHASE–PAS-DGC-EAL domain protein, in Azospirillum brasilense

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
José Francisco Cruz-Pérez ◽  
Roxana Lara-Oueilhe ◽  
Cynthia Marcos-Jiménez ◽  
Ricardo Cuatlayotl-Olarte ◽  
María Luisa Xiqui-Vázquez ◽  
...  
Keyword(s):  
Azospirillum Brasilense ◽  
Type Strain ◽  
Wild Type Strain ◽  
Soil Conditions ◽  
Wild Type ◽  
Gene Encoding ◽  
Wheat Roots ◽  
Genes Encoding ◽  
In The Wild ◽  
Plant Growth Promoting

AbstractThe plant growth-promoting bacterium Azospirillum brasilense contains several genes encoding proteins involved in the biosynthesis and degradation of the second messenger cyclic-di-GMP, which may control key bacterial functions, such as biofilm formation and motility. Here, we analysed the function and expression of the cdgD gene, encoding a multidomain protein that includes GGDEF-EAL domains and CHASE and PAS domains. An insertional cdgD gene mutant was constructed, and analysis of biofilm and extracellular polymeric substance production, as well as the motility phenotype indicated that cdgD encoded a functional diguanylate protein. These results were correlated with a reduced overall cellular concentration of cyclic-di-GMP in the mutant over 48 h compared with that observed in the wild-type strain, which was recovered in the complemented strain. In addition, cdgD gene expression was measured in cells growing under planktonic or biofilm conditions, and differential expression was observed when KNO3 or NH4Cl was added to the minimal medium as a nitrogen source. The transcriptional fusion of the cdgD promoter with the gene encoding the autofluorescent mCherry protein indicated that the cdgD gene was expressed both under abiotic conditions and in association with wheat roots. Reduced colonization of wheat roots was observed for the mutant compared with the wild-type strain grown in the same soil conditions. The Azospirillum-plant association begins with the motility of the bacterium towards the plant rhizosphere followed by the adsorption and adherence of these bacteria to plant roots. Therefore, it is important to study the genes that contribute to this initial interaction of the bacterium with its host plant.

Disruption of genes for the enhanced biosynthesis of α-ketoglutarate in Corynebacterium glutamicum

2012 ◽  
Vol 58 (3) ◽  
pp. 278-286 ◽  
Author(s):  
Jae-Hyung Jo ◽  
Hye-Young Seol ◽  
Yun-Bom Lee ◽  
Min-Hong Kim ◽  
Hyung-Hwan Hyun ◽  
...  
Keyword(s):  
Corynebacterium Glutamicum ◽  
Biosynthetic Pathway ◽  
Isocitrate Lyase ◽  
Glutamate Synthase ◽  
Flask Culture ◽  
Control Strain ◽  
Enhanced Production ◽  
Key Enzyme ◽  
Microbial Strains ◽  
Glyoxylate Pathway

The development of microbial strains for the enhanced production of α-ketoglutarate (α-KG) was investigated using a strain of Corynebacterium glutamicum that overproduces of l-glutamate, by disrupting three genes involved in the α-KG biosynthetic pathway. The pathways competing with the biosynthesis of α-KG were blocked by knocking out aceA (encoding isocitrate lyase, ICL), gdh (encoding glutamate dehydrogenase, l-gluDH), and gltB (encoding glutamate synthase or glutamate-2-oxoglutarate aminotransferase, GOGAT). The strain with aceA, gltB, and gdh disrupted showed reduced ICL activity and no GOGAT and l-gluDH activities, resulting in up to 16-fold more α-KG production than the control strain in flask culture. These results suggest that l-gluDH is the key enzyme in the conversion of α-KG to l-glutamate; therefore, prevention of this step could promote α-KG accumulation. The inactivation of ICL leads the carbon flow to α-KG by blocking the glyoxylate pathway. However, the disruption of gltB did not affect the biosynthesis of α-KG. Our results can be applied in the industrial production of α-KG by using C. glutamicum as producer.

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