Diverse allosteric and catalytic functions of tetrameric d-lactate dehydrogenases from three Gram-negative bacteria

Furukawa, Naomichi; Miyanaga, Akimasa; Togawa, M.; Nakajima, Masahiro; Taguchi, Hayao

doi:10.1186/s13568-014-0076-1

Cited by 17 publications

(47 citation statements)

References 41 publications

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“…In general, divalent metal ions have diverse effects on the catalytic activity of LDHs from different bacterial sources. In a study by Furukawa, Miyanaga, Togawa, Nakajima, and Taguchi (), Ca 2+ was shown to have a significant activation effect on Pseudomonas aeruginosa D‐LDH, whereas having no influence on Fusobacterium nucleatum D‐LDH and E. coli D‐LDH (). Furthermore, Mg 2+ activated F .…”

Section: Resultsmentioning

confidence: 99%

“…Furthermore, Mg 2+ activated F . nucleatum D‐LDH and P. aeruginosa D‐LDH, but was unable to activate E. coli D‐LDH (Furukawa et al., ). Jiang et al., reported that Ca 2+ activated B .…”

Section: Resultsmentioning

confidence: 99%

“…In contrast to D‐LDH1 and D‐LDH3, D‐LDH2 displayed a broad substrate specificity toward relative bulky 2‐ketocarboxylic acids, and exhibited the highest catalytic activity toward 2‐ketoisocaproate. The d ‐lactate dehydrogenases from Leuconostoc mesenteroides ATCC 8293, P. aeruginosa JCM8532 and L. jensenii SJ‐7A‐LIS were also reported to have narrow substrate specificities (Furukawa et al., ; Li et al., ), whereas d ‐lactate dehydrogenases from F. nucleatum ATCC25586 and L. jensenii 269‐3 exhibited relative broad substrate specificities (Furukawa et al., ; Jun et al., ). Among L‐LDHs, L‐LDH1 showed the highest catalytic activity toward pyruvate among the tested 2‐ketocarboxylic acids, whereas L‐LDH2 exhibited high activity to pyruvate analogs with larger groups, such as 2‐ketovalerate and 2‐ketoisocaproate.…”

Section: Resultsmentioning

confidence: 99%

“…Therefore, we constructed two phylogenetic trees to analyze evolutionary relationships of D‐LDHs and L‐LDHs from S. inulinus YBS1‐5, respectively. Overall, D‐LDHs clustered with a large D‐2‐hydroxyacid dehydrogenase superfamily, comprised of various kinds of D‐isomer dehydrogenases such as d ‐lactate dehydrogenase, D‐glycerate dehydrogenase, D‐phosphoglycerate dehydrogenase, D‐hydroxyglutarate dehydrogenase, D‐hydroxyisocaproate dehydrogenase, phosphite dehydrogenase, formate dehydrogenase, phenylpyruvate reductase, and 2‐ketopantoate reductase (Cristescu & Egbosima, ; Furukawa et al., ; Zhu et al., ). Although these enzymes possess highly divergent primary sequences, the core folding is structurally conserved, and the chiral product of the reaction is D‐isomer.…”

Section: Resultsmentioning

confidence: 99%

“…The substrate specificity of D-LDHs and L-LDHs was analyzed with a variety of 2-ketocarboxylic acids as presented in Figure 2b and c. D-LDH1 exhibited narrow substrate specificity toward pyruvate among 2-ketocarboxylic acid substrates, although it also showed a few catalytic activities toward glyoxylate and 2-ketobutyrate. (Furukawa et al, 2014;Jun et al, 2013). Among L-LDHs, L-LDH1…”

Section: Cloning Expression and Purification Of Ldhs From S Inulmentioning

confidence: 99%

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Relative catalytic efficiencies and transcript levels of three d‐ and two l‐lactate dehydrogenases for optically pure d‐lactate production in Sporolactobacillus inulinus

Zheng

et al. 2018

MicrobiologyOpen

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As the optical purity of the lactate monomer is pivotal for polymerization, the production of optically pure d-lactate is of significant importance. Sporolactobacillus inulinus YBS1-5 is a superior optically pure d-lactate-producing bacterium. However, little is known about the relationship between lactate dehydrogenases in S. inulinus YBS1-5 and the optical purity of d-lactate. Three potential d-lactate dehydrogenase (D-LDH1-3)- and two putative l-lactate dehydrogenase (L-LDH1-2)-encoding genes were cloned from the YBS1-5 strain and expressed in Escherichia coli D-LDH1 exhibited the highest catalytic efficiency toward pyruvate, whereas two L-LDHs showed low catalytic efficiency. Different neutralizers significantly affected the optical purity of d-lactate produced by strain YBS1-5 as well as the transcription levels of ldhDs and ldhLs. The high catalytic efficiency of D-LDH1 and elevated ldhD1 mRNA levels suggest that this enzyme is essential for d-lactate synthesis in S. inulinus YBS1-5. The correlation between the optical purity of d-lactate and transcription levels of ldhL1 in the case of different neutralizers indicate that ldhL1 is a key factor affecting the optical purity of d-lactate in S. inulinus YBS1-5.

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Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Section: Cloning Expression and Purification Of Ldhs From S Inulmentioning

confidence: 99%

See 3 more Smart Citations

Relative catalytic efficiencies and transcript levels of three d‐ and two l‐lactate dehydrogenases for optically pure d‐lactate production in Sporolactobacillus inulinus

Zheng

et al. 2018

MicrobiologyOpen

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Genetic tools for the redirection of the central carbon flow towards the production of lactate in the human gut bacterium Phocaeicola (Bacteroides) vulgatus

Lück

Deppenmeier

2022

Appl Microbiol Biotechnol

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Species of the genera Bacteroides and Phocaeicola play an important role in the human colon. The organisms contribute to the degradation of complex heteropolysaccharides to small chain fatty acids, which are in part utilized by the human body. Furthermore, these organisms are involved in the synthesis of vitamins and other bioactive compounds. Of special interest is Phocaeicola vulgatus, originally classified as a Bacteroides species, due to its abundance in the human intestinal tract and its ability to degrade many plant-derived heteropolysaccharides. We analyzed different tools for the genetic modification of this microorganism, with respect to homologous gene expression of the ldh gene encoding a D-lactate dehydrogenase (LDH). Therefore, the ldh gene was cloned into the integration vector pMM656 and the shuttle vector pG106 for homologous gene expression in P. vulgatus. We determined the ldh copy number, transcript abundance, and the enzyme activity of the wild type and the mutants. The strain containing the shuttle vector showed an approx. 1500-fold increase in the ldh transcript concentration and an enhanced LDH activity that was about 200-fold higher compared to the parental strain. Overall, the proportion of lactate in the general catabolic carbon flow increased from 2.9% (wild type) to 28.5% in the LDH-overproducing mutant. This approach is a proof of concept, verifying the genetic accessibility of P. vulgatus and could form the basis for targeted genetic optimization. Key points • A lactate dehydrogenase was overexpressed in Phocaeicola (Bacteroides) vulgatus. • The ldh transcript abundance and the LDH activity increased sharply in the mutant. • The proportion of lactate in the catabolic carbon flow increased to about 30%.

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A synthetic pathway for the production of 2-hydroxyisovaleric acid in Escherichia coli

Cheong

Clomburg

González

2018

Journal of Industrial Microbiology and Biotechnology

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Synthetic biology, encompassing the design and construction of novel artificial biological pathways and organisms and the redesign of existing natural biological systems, is rapidly expanding the number of applications for which biological systems can play an integral role. In the context of chemical production, the combination of synthetic biology and metabolic engineering approaches continues to unlock the ability to biologically produce novel and complex molecules from a variety of feedstocks. Here, we utilize a synthetic approach to design and build a pathway to produce 2-hydroxyisovaleric acid in Escherichia coli and demonstrate how pathway design can be supplemented with metabolic engineering approaches to improve pathway performance from various carbon sources. Drawing inspiration from the native pathway for the synthesis of the 5-carbon amino acid L-valine, we exploit the decarboxylative condensation of two molecules of pyruvate, with subsequent reduction and dehydration reactions enabling the synthesis of 2-hydroxyisovaleric acid. Key to our approach was the utilization of an acetolactate synthase which minimized kinetic and regulatory constraints to ensure sufficient flux entering the pathway. Critical host modifications enabling maximum product synthesis from either glycerol or glucose were then examined, with the varying degree of reduction of these carbons sources playing a major role in the required host background. Through these engineering efforts, the designed pathway produced 6.2 g/L 2-hydroxyisovaleric acid from glycerol at 58% of maximum theoretical yield and 7.8 g/L 2-hydroxyisovaleric acid from glucose at 73% of maximum theoretical yield. These results demonstrate how the combination of synthetic biology and metabolic engineering approaches can facilitate bio-based chemical production.

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Diverse allosteric and catalytic functions of tetrameric d-lactate dehydrogenases from three Gram-negative bacteria

Cited by 17 publications

References 41 publications

Relative catalytic efficiencies and transcript levels of three d‐ and two l‐lactate dehydrogenases for optically pure d‐lactate production in Sporolactobacillus inulinus

Relative catalytic efficiencies and transcript levels of three d‐ and two l‐lactate dehydrogenases for optically pure d‐lactate production in Sporolactobacillus inulinus

Genetic tools for the redirection of the central carbon flow towards the production of lactate in the human gut bacterium Phocaeicola (Bacteroides) vulgatus

A synthetic pathway for the production of 2-hydroxyisovaleric acid in Escherichia coli

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