PUMA publications for /tag/Dehydrogenase,https://puma.ub.uni-stuttgart.de/tag/Dehydrogenase,PUMA RSS feed for /tag/Dehydrogenase,2024-03-28T11:59:40+01:00Current knowledge on isobutanol production with Escherichia coli, Bacillus subtilis and Corynebacterium glutamicumhttps://puma.ub.uni-stuttgart.de/bibtex/20f88c2605071e1c350ed8a296eece367/bastianbastian2018-02-09T13:18:17+01:00Acids, Alcohol Bacillus Bacterial Butanols, Carboxy-Lyases, Corynebacterium Dehydrogenase, Engineering Escherichia Industrial Keto Metabolic Microbiology, Proteins, Recombinant coli, glutamicum, myown subtilis, <span data-person-type="author" class="authorEditorList "><span><span itemtype="http://schema.org/Person" itemscope="itemscope" itemprop="author"><a title="Bastian Blombach" itemprop="url" href="/person/133960b6afe49761f6398812d65d1660e/author/0"><span itemprop="name">B. Blombach</span></a></span>, </span> and <span><span itemtype="http://schema.org/Person" itemscope="itemscope" itemprop="author"><a title="Bernhard J. Eikmanns" itemprop="url" href="/person/133960b6afe49761f6398812d65d1660e/author/1"><span itemprop="name">B. Eikmanns</span></a></span></span>. </span><span class="additional-entrytype-information"><span itemtype="http://schema.org/PublicationIssue" itemscope="itemscope" itemprop="isPartOf"><em><span itemprop="journal">Bioeng Bugs</span>, </em> <em><span itemtype="http://schema.org/PublicationVolume" itemscope="itemscope" itemprop="isPartOf"><span itemprop="volumeNumber">2 </span></span>(<span itemprop="issueNumber">6</span>):
<span itemprop="pagination">346--350</span></em> </span>(<em><span>December 2011<meta content="December 2011" itemprop="datePublished"/></span></em>)</span>Fri Feb 09 13:18:17 CET 2018Bioeng Bugsdec6346--350Current knowledge on isobutanol production with {Escherichia} coli, {Bacillus} subtilis and {Corynebacterium} glutamicum22011Acids, Alcohol Bacillus Bacterial Butanols, Carboxy-Lyases, Corynebacterium Dehydrogenase, Engineering Escherichia Industrial Keto Metabolic Microbiology, Proteins, Recombinant coli, glutamicum, myown subtilis, Due to steadily rising crude oil prices great efforts have been made to develop designer bugs for the fermentative production of higher alcohols, such as 2-methyl-1-butanol, 3-methyl-1-butanol and 2-Methyl-1-propanol (isobutanol), which all possess quality characteristics comparable to traditional oil based fuels. The common metabolic engineering approach uses the last two steps of the Ehrlich pathway, catalyzed by 2-ketoacid decarboxylase and an alcohol dehydrogenase converting the branched chain 2-ketoacids of L-isoleucine, L-leucine, and L-valine into the respective alcohols. This strategy was successfully used to engineer well suited and industrially employed bacteria, such as Escherichia coli, Bacillus subtilis and Corynebacterium glutamicum for the production of higher alcohols. Among these alcohols, isobutanol is currently the most promising one regarding final titer and yield. This article summarizes the current knowledge and achievements on isobutanol production with E. coli, B. subtilis and C. glutamicum regarding the metabolic engineering approaches and process conditions.Stereospecificity of Corynebacterium glutamicum 2,3-butanediol dehydrogenase and implications for the stereochemical purity of bioproduced 2,3-butanediolhttps://puma.ub.uni-stuttgart.de/bibtex/2d2fbd13b8243050d76f6784383286149/bastianbastian2018-02-09T13:18:17+01:002,3-Butanediol, Acetoin, Acetolactate Alcohol Butanediol Butylene Carboxy-Lyases, Corynebacterium Engineering Escherichia Glycols, Lactococcus Magnetic Metabolic Oxidoreductases, Proteins, Recombinant Resonance Specificity, Spectroscopy, Stereospecificity, Substrate Synthase, coli, dehydrogenase, glutamicum, lactis, myown <span data-person-type="author" class="authorEditorList "><span><span itemtype="http://schema.org/Person" itemscope="itemscope" itemprop="author"><a title="Dušica Radoš" itemprop="url" href="/person/1dd4270d58d31233ae25eebd4b8cf903e/author/0"><span itemprop="name">D. Radoš</span></a></span>, </span><span><span itemtype="http://schema.org/Person" itemscope="itemscope" itemprop="author"><a title="David L. Turner" itemprop="url" href="/person/1dd4270d58d31233ae25eebd4b8cf903e/author/1"><span itemprop="name">D. Turner</span></a></span>, </span><span><span itemtype="http://schema.org/Person" itemscope="itemscope" itemprop="author"><a title="Teresa Catarino" itemprop="url" href="/person/1dd4270d58d31233ae25eebd4b8cf903e/author/2"><span itemprop="name">T. Catarino</span></a></span>, </span><span><span itemtype="http://schema.org/Person" itemscope="itemscope" itemprop="author"><a title="Eugenia Hoffart" itemprop="url" href="/person/1dd4270d58d31233ae25eebd4b8cf903e/author/3"><span itemprop="name">E. Hoffart</span></a></span>, </span><span><span itemtype="http://schema.org/Person" itemscope="itemscope" itemprop="author"><a title="Ana Rute Neves" itemprop="url" href="/person/1dd4270d58d31233ae25eebd4b8cf903e/author/4"><span itemprop="name">A. Neves</span></a></span>, </span><span><span itemtype="http://schema.org/Person" itemscope="itemscope" itemprop="author"><a title="Bernhard J. Eikmanns" itemprop="url" href="/person/1dd4270d58d31233ae25eebd4b8cf903e/author/5"><span itemprop="name">B. Eikmanns</span></a></span>, </span><span><span itemtype="http://schema.org/Person" itemscope="itemscope" itemprop="author"><a title="Bastian Blombach" itemprop="url" href="/person/1dd4270d58d31233ae25eebd4b8cf903e/author/6"><span itemprop="name">B. Blombach</span></a></span>, </span> and <span><span itemtype="http://schema.org/Person" itemscope="itemscope" itemprop="author"><a title="Helena Santos" itemprop="url" href="/person/1dd4270d58d31233ae25eebd4b8cf903e/author/7"><span itemprop="name">H. Santos</span></a></span></span>. </span><span class="additional-entrytype-information"><span itemtype="http://schema.org/PublicationIssue" itemscope="itemscope" itemprop="isPartOf"><em><span itemprop="journal">Appl. Microbiol. Biotechnol.</span>, </em> <em><span itemtype="http://schema.org/PublicationVolume" itemscope="itemscope" itemprop="isPartOf"><span itemprop="volumeNumber">100 </span></span>(<span itemprop="issueNumber">24</span>):
<span itemprop="pagination">10573--10583</span></em> </span>(<em><span>December 2016<meta content="December 2016" itemprop="datePublished"/></span></em>)</span>Fri Feb 09 13:18:17 CET 2018Appl. Microbiol. Biotechnol.dec2410573--10583Stereospecificity of {Corynebacterium} glutamicum 2,3-butanediol dehydrogenase and implications for the stereochemical purity of bioproduced 2,3-butanediol10020162,3-Butanediol, Acetoin, Acetolactate Alcohol Butanediol Butylene Carboxy-Lyases, Corynebacterium Engineering Escherichia Glycols, Lactococcus Magnetic Metabolic Oxidoreductases, Proteins, Recombinant Resonance Specificity, Spectroscopy, Stereospecificity, Substrate Synthase, coli, dehydrogenase, glutamicum, lactis, myown The stereochemistry of 2,3-butanediol (2,3-BD) synthesis in microbial fermentations is important for many applications. In this work, we showed that Corynebacterium glutamicum endowed with the Lactococcus lactis genes encoding α-acetolactate synthase and decarboxylase activities produced meso-2,3-BD as the major end product, meaning that (R)-acetoin is a substrate for endogenous 2,3-butanediol dehydrogenase (BDH) activity. This is curious in view of the reported absolute stereospecificity of C. glutamicum BDH for (S)-acetoin (Takusagawa et al. Biosc Biotechnol Biochem 65:1876-1878, 2001). To resolve this discrepancy, the enzyme encoded by butA Cg was produced in Escherichia coli and purified, and the stereospecific properties of the pure protein were examined. Activity assays monitored online by 1H-NMR using racemic acetoin and an excess of NADH showed an initial, fast production of (2S,3S)-2,3-BD, followed by a slow (∼20-fold lower apparent rate) formation of meso-2,3-BD. Kinetic parameters for (S)-acetoin, (R)-acetoin, meso-2,3-BD and (2S,3S)-BD were determined by spectrophotometric assays. V max values for (S)-acetoin and (R)-acetoin were 119 ± 15 and 5.23 ± 0.06 μmol min-1 mg protein-1, and K m values were 0.23 ± 0.02 and 1.49 ± 0.07 mM, respectively. We conclude that C. glutamicum BDH is not absolutely specific for (S)-acetoin, though this is the preferred substrate. Importantly, the low activity of BDH with (R)-acetoin was sufficient to support high yields of meso-2,3-BD in the engineered strain C. glutamicum ΔaceEΔpqoΔldhA(pEKEx2-als,aldB,butA Cg ). Additionally, we found that the BDH activity was nearly abolished upon inactivation of butA Cg (from 0.30 ± 0.03 to 0.004 ± 0.001 μmol min-1 mg protein-1), indicating that C. glutamicum expresses a single BDH under the experimental conditions examined.Engineering Corynebacterium glutamicum for the production of 2,3-butanediolhttps://puma.ub.uni-stuttgart.de/bibtex/29ce34124eaffe2a799aade38deec413d/bastianbastian2018-02-09T13:18:17+01:00Bacterial Bioreactors, Butylene Complex, Corynebacterium Dehydrogenase Dehydrogenase, Engineering Family, Glucose, Glycols, L-Lactate Lactococcus Metabolic Multigene Oxygen, Proteins, Pyruvate glutamicum, lactis, myown <span data-person-type="author" class="authorEditorList "><span><span itemtype="http://schema.org/Person" itemscope="itemscope" itemprop="author"><a title="Dušica Radoš" itemprop="url" href="/person/1518ec5750d920964df1a659788edff11/author/0"><span itemprop="name">D. Radoš</span></a></span>, </span><span><span itemtype="http://schema.org/Person" itemscope="itemscope" itemprop="author"><a title="Ana Lúcia Carvalho" itemprop="url" href="/person/1518ec5750d920964df1a659788edff11/author/1"><span itemprop="name">A. Carvalho</span></a></span>, </span><span><span itemtype="http://schema.org/Person" itemscope="itemscope" itemprop="author"><a title="Stefan Wieschalka" itemprop="url" href="/person/1518ec5750d920964df1a659788edff11/author/2"><span itemprop="name">S. Wieschalka</span></a></span>, </span><span><span itemtype="http://schema.org/Person" itemscope="itemscope" itemprop="author"><a title="Ana Rute Neves" itemprop="url" href="/person/1518ec5750d920964df1a659788edff11/author/3"><span itemprop="name">A. Neves</span></a></span>, </span><span><span itemtype="http://schema.org/Person" itemscope="itemscope" itemprop="author"><a title="Bastian Blombach" itemprop="url" href="/person/1518ec5750d920964df1a659788edff11/author/4"><span itemprop="name">B. Blombach</span></a></span>, </span><span><span itemtype="http://schema.org/Person" itemscope="itemscope" itemprop="author"><a title="Bernhard J. Eikmanns" itemprop="url" href="/person/1518ec5750d920964df1a659788edff11/author/5"><span itemprop="name">B. Eikmanns</span></a></span>, </span> and <span><span itemtype="http://schema.org/Person" itemscope="itemscope" itemprop="author"><a title="Helena Santos" itemprop="url" href="/person/1518ec5750d920964df1a659788edff11/author/6"><span itemprop="name">H. Santos</span></a></span></span>. </span><span class="additional-entrytype-information"><span itemtype="http://schema.org/PublicationIssue" itemscope="itemscope" itemprop="isPartOf"><em><span itemprop="journal">Microb. Cell Fact.</span>, </em> </span>(<em><span>October 2015<meta content="October 2015" itemprop="datePublished"/></span></em>)</span>Fri Feb 09 13:18:17 CET 2018Microb. Cell Fact.oct171Engineering {Corynebacterium} glutamicum for the production of 2,3-butanediol142015Bacterial Bioreactors, Butylene Complex, Corynebacterium Dehydrogenase Dehydrogenase, Engineering Family, Glucose, Glycols, L-Lactate Lactococcus Metabolic Multigene Oxygen, Proteins, Pyruvate glutamicum, lactis, myown BACKGROUND: 2,3-Butanediol is an important bulk chemical with a wide range of applications. In bacteria, this metabolite is synthesised from pyruvate via a three-step pathway involving α-acetolactate synthase, α-acetolactate decarboxylase and 2,3-butanediol dehydrogenase. Thus far, the best producers of 2,3-butanediol are pathogenic strains, hence, the development of more suitable organisms for industrial scale fermentation is needed. Herein, 2,3-butanediol production was engineered in the Generally Regarded As Safe (GRAS) organism Corynebacterium glutamicum. A two-stage fermentation process was implemented: first, cells were grown aerobically on acetate; in the subsequent production stage cells were used to convert glucose into 2,3-butanediol under non-growing and oxygen-limiting conditions.
RESULTS: A gene cluster, encoding the 2,3-butanediol biosynthetic pathway of Lactococcus lactis, was assembled and expressed in background strains, C. glutamicum ΔldhA, C. glutamicum ΔaceEΔpqoΔldhA and C. glutamicum ΔaceEΔpqoΔldhAΔmdh, tailored to minimize pyruvate-consuming reactions, i.e., to prevent carbon loss in lactic, acetic and succinic acids. Producer strains were characterized in terms of activity of the relevant enzymes in the 2,3-butanediol forming pathway, growth, and production of 2,3-butanediol under oxygen-limited conditions. Productivity was maximized by manipulating the aeration rate in the production phase. The final strain, C. glutamicum ΔaceEΔpqoΔldhAΔmdh(pEKEx2-als,aldB,Ptuf butA), under optimized conditions produced 2,3-butanediol with a 0.66 mol mol(-1) yield on glucose, an overall productivity of 0.2 g L(-1) h(-1) and a titer of 6.3 g L(-1).
CONCLUSIONS: We have successfully developed C. glutamicum into an efficient cell factory for 2,3-butanediol production. The use of the engineered strains as a basis for production of acetoin, a widespread food flavour, is proposed.Impact of different CO2/HCO3- levels on metabolism and regulation in Corynebacterium glutamicumhttps://puma.ub.uni-stuttgart.de/bibtex/24eb5c8cfb1d770de7b680bff723465f7/bastianbastian2018-02-09T13:18:17+01:00Alanine, Bacterial Bacterial, Biomass, CO(2)/HCO(3)(−), Carbon Corynebacterium DNA-Binding Dehydrogenase, Dioxide, Diphtheria Dissolved DtxR, Expression Gene Glucose, Glucosephosphate Proteins, Regulation, Thiamin Thiamine, Toxin, Valine, biosynthesis carbon dioxide, glutamicum, myown repressor toxin <span data-person-type="author" class="authorEditorList "><span><span itemtype="http://schema.org/Person" itemscope="itemscope" itemprop="author"><a title="Bastian Blombach" itemprop="url" href="/person/175535e85947aaa1c453c03c26dfd2dfc/author/0"><span itemprop="name">B. Blombach</span></a></span>, </span><span><span itemtype="http://schema.org/Person" itemscope="itemscope" itemprop="author"><a title="Jens Buchholz" itemprop="url" href="/person/175535e85947aaa1c453c03c26dfd2dfc/author/1"><span itemprop="name">J. Buchholz</span></a></span>, </span><span><span itemtype="http://schema.org/Person" itemscope="itemscope" itemprop="author"><a title="Tobias Busche" itemprop="url" href="/person/175535e85947aaa1c453c03c26dfd2dfc/author/2"><span itemprop="name">T. Busche</span></a></span>, </span><span><span itemtype="http://schema.org/Person" itemscope="itemscope" itemprop="author"><a title="Jörn Kalinowski" itemprop="url" href="/person/175535e85947aaa1c453c03c26dfd2dfc/author/3"><span itemprop="name">J. Kalinowski</span></a></span>, </span> and <span><span itemtype="http://schema.org/Person" itemscope="itemscope" itemprop="author"><a title="Ralf Takors" itemprop="url" href="/person/175535e85947aaa1c453c03c26dfd2dfc/author/4"><span itemprop="name">R. Takors</span></a></span></span>. </span><span class="additional-entrytype-information"><span itemtype="http://schema.org/PublicationIssue" itemscope="itemscope" itemprop="isPartOf"><em><span itemprop="journal">J. Biotechnol.</span>, </em> <em><span itemtype="http://schema.org/PublicationVolume" itemscope="itemscope" itemprop="isPartOf"><span itemprop="volumeNumber">168 </span></span>(<span itemprop="issueNumber">4</span>):
<span itemprop="pagination">331--340</span></em> </span>(<em><span>December 2013<meta content="December 2013" itemprop="datePublished"/></span></em>)</span>Fri Feb 09 13:18:17 CET 2018J. Biotechnol.dec4331--340Impact of different {CO}2/{HCO}3- levels on metabolism and regulation in {Corynebacterium} glutamicum1682013Alanine, Bacterial Bacterial, Biomass, CO(2)/HCO(3)(−), Carbon Corynebacterium DNA-Binding Dehydrogenase, Dioxide, Diphtheria Dissolved DtxR, Expression Gene Glucose, Glucosephosphate Proteins, Regulation, Thiamin Thiamine, Toxin, Valine, biosynthesis carbon dioxide, glutamicum, myown repressor toxin We investigated the growth kinetics and transcriptional responses of Corynebacterium glutamicum in environments with low (pCO2{\textless}40 mbar) and high (pCO2 ≥ 300 mbar) CO2/HCO3(-) levels compared to standard conditions. When cultivated at high CO2/HCO3(-)-levels, C. glutamicum showed increased (63\%) biomass to substrate yields during the initial growth phase. Other kinetic parameters such as growth rate (μ), specific glucose consumption rate (qS), and selected enzymatic activities of anaplerotic reactions, the pentose phosphate pathway and the tricarboxylic acid cycle were similar to standard conditions. However, microarray hybridization disclosed a complex transcriptional response involving 117 differentially expressed genes. Among those, 60 genes were assigned to the complete DtxR/RipA regulon controlling iron homeostasis in C. glutamicum. Impaired growth of a ΔdtxR mutant at high CO2/HCO3(-) levels validated the relevance of this master regulator to cope with excessive CO2/HCO3(-) availability. At low CO2/HCO3(-) levels, C. glutamicum grew in a bi-level manner with three distinct growth phases. Differential analyses revealed approximately doubled activities of glucose-6-phosphate dehydrogenase and 6-phosphogluconate dehydrogenase accompanied by the formation of L-alanine and L-valine during the lowest μ occurring in mid-phase of the cultivation. DNA microarray analysis revealed more than 100 differentially expressed genes in growth phase II compared to phase I including almost all thiamin pyrophosphate (TPP) biosynthesis genes, which were significantly up regulated. Concluding, we hypothesize that C. glutamicum counteracts the lack of CO2/HCO3(-) by triggering TPP biosynthesis for increasing the activities of TPP-dependent enzymes involved in CO2 formation.