Chirality Matters: Synthesis and Consumption of the d-Enantiomer of Lactic Acid by Synechocystis sp. Strain PCC6803

ABSTRACTBoth enantiomers of lactic acid,l-lactic acid andd-lactic acid, can be produced in a sustainable way by a photosynthetic microbial cell factory and thus from CO2, sunlight, and water. Several properties of polylactic acid (a polyester of polymerized lactic acid) depend on the controlled blend of these two enantiomers. Recently, cyanobacteriumSynechocystissp. strain PCC6803 was genetically modified to allow formation of either of these two enantiomers. This report elaborates on thed-lactic acid production achieved by the introduction of ad-specific lactate dehydrogenase from the lactic acid bacteriumLeuconostoc mesenteroidesintoSynechocystis. A typical batch culture of this recombinant strain initially shows lactic acid production, followed by a phase of lactic acid consumption, until production “outcompetes” consumption at later growth stages. We show thatSynechocystisis able to used-lactic acid, but notl-lactic acid, as a carbon source for growth. Deletion of the organism's putatived-lactate dehydrogenase (encoded byslr1556), however, does not eliminate this ability with respect tod-lactic acid consumption. In contrast,d-lactic acid consumption does depend on the presence of glycolate dehydrogenase GlcD1 (encoded bysll0404). Accordingly, this report highlights the need to match a product of interest of a cyanobacterial cell factory with the metabolic network present in the host used for its synthesis and emphasizes the need to understand the physiology of the production host in detail.

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Major Role of NAD-Dependent Lactate Dehydrogenases in the Production of l-Lactic Acid with High Optical Purity by the Thermophile Bacillus coagulans

Applied and Environmental Microbiology ◽

10.1128/aem.01864-14 ◽

2014 ◽

Vol 80 (23) ◽

pp. 7134-7141 ◽

Cited By ~ 26

Author(s):

Limin Wang ◽

Yumeng Cai ◽

Lingfeng Zhu ◽

Honglian Guo ◽

Bo Yu

Keyword(s):

Lactic Acid ◽

Lactate Dehydrogenase ◽

Acid Production ◽

Lactic Acid Production ◽

Optical Purity ◽

Bacillus Coagulans ◽

Lactobacillus Delbrueckii ◽

Content Type ◽

High Optical Purity ◽

Optically Pure

ABSTRACTBacillus coagulans2-6 is an excellent producer of optically purel-lactic acid. However, little is known about the mechanism of synthesis of the highly optically purel-lactic acid produced by this strain. Three enzymes responsible for lactic acid production—NAD-dependentl-lactate dehydrogenase (l-nLDH; encoded byldhL), NAD-dependentd-lactate dehydrogenase (d-nLDH; encoded byldhD), and glycolate oxidase (GOX)—were systematically investigated in order to study the relationship between these enzymes and the optical purity of lactic acid.Lactobacillus delbrueckiisubsp.bulgaricusDSM 20081 (ad-lactic acid producer) andLactobacillus plantarumsubsp.plantarumDSM 20174 (adl-lactic acid producer) were also examined in this study as comparative strains, in addition toB. coagulans. The specific activities of key enzymes for lactic acid production in the three strains were characterizedin vivoandin vitro, and the levels of transcription of theldhL,ldhD, and GOX genes during fermentation were also analyzed. The catalytic activities ofl-nLDH andd-nLDH were different inl-,d-, anddl-lactic acid producers. Onlyl-nLDH activity was detected inB. coagulans2-6 under native conditions, and the level of transcription ofldhLinB. coagulans2-6 was much higher than that ofldhDor the GOX gene at all growth phases. However, for the twoLactobacillusstrains used in this study,ldhDtranscription levels were higher than those ofldhL. The high catalytic efficiency ofl-nLDH toward pyruvate and the high transcription ratios ofldhLtoldhDandldhLto the GOX gene provide the key explanations for the high optical purity ofl-lactic acid produced byB. coagulans2-6.

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Engineering a Cyanobacterial Cell Factory for Production of Lactic Acid

Applied and Environmental Microbiology ◽

10.1128/aem.01587-12 ◽

2012 ◽

Vol 78 (19) ◽

pp. 7098-7106 ◽

Cited By ~ 126

Author(s):

S. Andreas Angermayr ◽

Michal Paszota ◽

Klaas J. Hellingwerf

Keyword(s):

Lactic Acid ◽

Lactate Dehydrogenase ◽

Lactic Acid Production ◽

Lactate Production ◽

Cell Factory ◽

Wild Type ◽

Extracellular Medium ◽

Content Type ◽

Bulk Chemicals ◽

Versatile Tool

ABSTRACTMetabolic engineering of microorganisms has become a versatile tool to facilitate production of bulk chemicals, fuels, etc. Accordingly, CO2has been exploited via cyanobacterial metabolism as a sustainable carbon source of biofuel and bioplastic precursors. Here we extended these observations by showing that integration of anldhgene fromBacillus subtilis(encoding anl-lactate dehydrogenase) into the genome ofSynechocystissp. strain PCC6803 leads tol-lactic acid production, a phenotype which is shown to be stable for prolonged batch culturing. Coexpression of a heterologous soluble transhydrogenase leads to an even higher lactate production rate and yield (lactic acid accumulating up to a several-millimolar concentration in the extracellular medium) than those for the singleldhmutant. The expression of a transhydrogenase alone, however, appears to be harmful to the cells, and a mutant carrying such a gene is rapidly outcompeted by a revertant(s) with a wild-type growth phenotype. Furthermore, our results indicate that the introduction of a lactate dehydrogenase rescues this phenotype by preventing the reversion.

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Production of Raw Starch-Digesting Amylolytic Preparation in Yarrowia lipolytica and Its Application in Biotechnological Synthesis of Lactic Acid and Ethanol

Microorganisms ◽

10.3390/microorganisms8050717 ◽

2020 ◽

Vol 8 (5) ◽

pp. 717

Author(s):

Aleksandra Gęsicka ◽

Monika Borkowska ◽

Wojciech Białas ◽

Paulina Kaczmarek ◽

Ewelina Celińska

Keyword(s):

Lactic Acid ◽

Ethanol Production ◽

Yarrowia Lipolytica ◽

Acid Production ◽

Lactic Acid Production ◽

Thermal Conditions ◽

Cell Factory ◽

The Novel ◽

Microbial Cell Factory ◽

Raw Starch

Sustainable economy drives increasing demand for raw biomass-decomposing enzymes. Microbial expression platforms exploited as cellular factories of such biocatalysts meet requirements of large-volume production. Previously, we developed Yarrowia lipolytica recombinant strains able to grow on raw starch of different plant origin. In the present study, we used the most efficient amylolytic strain as a microbial cell factory of raw-starch-digesting (RSD) amylolytic preparation composed of two enzymes. The RSD-preparation was produced in fed-batch bioreactor cultures. Concentrated and partly purified preparation was then tested in simultaneous saccharification and fermentation (SSF) processes with thermotolerant Kluyveromyces marxianus for ethanol production and Lactobacillus plantarum for production of lactic acid. These processes were conducted as a proof-of-concept that application of the novel RSD-preparation supports sufficient starch hydrolysis enabling microbial growth and production of targeted molecules, as the selected strains were confirmed to lack amylolytic activity. Doses of the preparation and thermal conditions were individually adjusted for the two processes. Additionally, ethanol production was tested under different aeration strategies; and lactic acid production process was tested in thermally pre-treated substrate, as well. Conducted studies demonstrated that the novel RSD-preparation provides satisfactory starch hydrolyzing activity for ethanol and lactic acid production from starch by non-amylolytic microorganisms.

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Cloning of d-lactate dehydrogenase genes of Lactobacillus delbrueckii subsp. bulgaricus and their roles in d-lactic acid production

3 Biotech ◽

10.1007/s13205-017-0822-6 ◽

2017 ◽

Vol 7 (3) ◽

Cited By ~ 1

Author(s):

Yanna Huang ◽

Chunping You ◽

Zhenmin Liu

Keyword(s):

Lactic Acid ◽

Lactate Dehydrogenase ◽

Acid Production ◽

Lactic Acid Production ◽

Lactobacillus Delbrueckii ◽

Lactobacillus Delbrueckii Subsp

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Production of biopolymer precursors beta-alanine and L-lactic acid from CO2 with metabolically versatile Rhodococcus opacus DSM 43205

10.1101/2020.08.26.267856 ◽

2020 ◽

Author(s):

Laura Salusjärvi ◽

Leo Ojala ◽

Gopal Peddinti ◽

Michael Lienemann ◽

Paula Jouhten ◽

...

Keyword(s):

Lactic Acid ◽

Metabolic Engineering ◽

Lactate Dehydrogenase ◽

Acid Production ◽

Lactic Acid Production ◽

Water Electrolysis ◽

Rhodococcus Opacus ◽

Beta Alanine ◽

Polymer Precursors ◽

Autotrophic Bacteria

AbstractHydrogen oxidizing autotrophic bacteria are promising hosts for CO2 conversion into chemicals. In this work, we engineered the metabolically versatile lithoautotrophic bacterium Rhodococcus opacus strain DSM 43205 for synthesis of polymer precursors. Aspartate decarboxylase (panD) or lactate dehydrogenase (ldh) were expressed for beta-alanine or L-lactic acid production, respectively. The heterotrophic cultivations on glucose produced 25 mg L-1 beta-alanine and 742 mg L-1 L-lactic acid, while autotrophic cultivations with CO2, H2 and O2 resulted in the production of 1.8 mg L-1 beta-alanine and 146 mg L-1 L-lactic acid. Beta-alanine was also produced at 345 µg L-1 from CO2 in electrobioreactors, where H2 and O2 were provided by water electrolysis. This work demonstrates that R. opacus DSM 43205 can be readily engineered to produce chemicals from CO2 and provides base for its further metabolic engineering.

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Plasticity of the Pyruvate Node Modulates Hydrogen Peroxide Production and Acid Tolerance in Multiple Oral Streptococci

Applied and Environmental Microbiology ◽

10.1128/aem.01697-17 ◽

2017 ◽

Vol 84 (2) ◽

Cited By ~ 8

Author(s):

Xingqun Cheng ◽

Sylvio Redanz ◽

Nyssa Cullin ◽

Xuedong Zhou ◽

Xin Xu ◽

...

Keyword(s):

Lactic Acid ◽

Environmental Conditions ◽

Dental Plaque ◽

Acid Production ◽

Lactic Acid Production ◽

Acid Tolerance ◽

Oral Bacteria ◽

Low Ph ◽

Content Type ◽

Pyruvate Node

ABSTRACTCommensalStreptococcus sanguinisandStreptococcus gordoniiare pioneer oral biofilm colonizers. Characteristic for both is the SpxB-dependent production of H2O2, which is crucial for inhibiting competing biofilm members, especially the cariogenic speciesStreptococcus mutans. H2O2production is strongly affected by environmental conditions, but few mechanisms are known. Dental plaque pH is one of the key parameters dictating dental plaque ecology and ultimately oral health status. Therefore, the objective of the current study was to characterize the effects of environmental pH on H2O2production byS. sanguinisandS. gordonii.S. sanguinisH2O2production was not found to be affected by moderate changes in environmental pH, whereasS. gordoniiH2O2production declined markedly in response to lower pH. Further investigation into the pyruvate node, the central metabolic switch modulating H2O2or lactic acid production, revealed increased lactic acid levels forS. gordoniiat pH 6. The bias for lactic acid production at pH 6 resulted in concomitant improvement in the survival ofS. gordoniiat low pH and seems to constitute part of the acid tolerance response ofS. gordonii. Differential responses to pH similarly affect other oral streptococcal species, suggesting that the observed results are part of a larger phenomenon linking environmental pH, central metabolism, and the capacity to produce antagonistic amounts of H2O2.IMPORTANCEOral biofilms are subject to frequent and dramatic changes in pH.S. sanguinisandS. gordoniican compete with caries- and periodontitis-associated pathogens by generating H2O2. Therefore, it is crucial to understand howS. sanguinisandS. gordoniiadapt to low pH and maintain their competitiveness under acid stress. The present study provides evidence that certain oral bacteria respond to environmental pH changes by tuning their metabolic output in favor of lactic acid production, to increase their acid survival, while others maintain their H2O2production at a constant level. The differential control of H2O2production provides important insights into the role of environmental conditions for growth competition of the oral flora.

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Efficient Production of l-Lactic Acid by Metabolically Engineered Saccharomyces cerevisiae with a Genome-Integrated l-Lactate Dehydrogenase Gene

Applied and Environmental Microbiology ◽

10.1128/aem.71.4.1964-1970.2005 ◽

2005 ◽

Vol 71 (4) ◽

pp. 1964-1970 ◽

Cited By ~ 93

Author(s):

Nobuhiro Ishida ◽

Satoshi Saitoh ◽

Kenro Tokuhiro ◽

Eiji Nagamori ◽

Takashi Matsuyama ◽

...

Keyword(s):

Lactic Acid ◽

Lactate Dehydrogenase ◽

Acid Production ◽

Lactic Acid Production ◽

Bifidobacterium Longum ◽

Yeast Cells ◽

Efficient Production ◽

Coding Region ◽

A Genome ◽

Dehydrogenase Gene

ABSTRACT We developed a metabolically engineered yeast which produces lactic acid efficiently. In this recombinant strain, the coding region for pyruvate decarboxylase 1 (PDC1) on chromosome XII is substituted for that of the l-lactate dehydrogenase gene (LDH) through homologous recombination. The expression of mRNA for the genome-integrated LDH is regulated under the control of the native PDC1 promoter, while PDC1 is completely disrupted. Using this method, we constructed a diploid yeast transformant, with each haploid genome having a single insertion of bovine LDH. Yeast cells expressing LDH were observed to convert glucose to both lactate (55.6 g/liter) and ethanol (16.9 g/liter), with up to 62.2% of the glucose being transformed into lactic acid under neutralizing conditions. This transgenic strain, which expresses bovine LDH under the control of the PDC1 promoter, also showed high lactic acid production (50.2 g/liter) under nonneutralizing conditions. The differences in lactic acid production were compared among four different recombinants expressing a heterologous LDH gene (i.e., either the bovine LDH gene or the Bifidobacterium longum LDH gene): two transgenic strains with 2μm plasmid-based vectors and two genome-integrated strains.

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