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engineering corynebacterium glutamicum for the production of pyruvate.a corynebacterium glutamicum strain with inactivated pyruvate dehydrogenase complex and a deletion of the gene encoding the pyruvate:quinone oxidoreductase produces about 19 mm l: -valine, 28 mm l: -alanine and about 55 mm pyruvate from 150 mm glucose. based on this double mutant c. glutamicum △acee △pqo, we engineered c. glutamicum for efficient production of pyruvate from glucose by additional deletion of the ldha gene encoding nad(+)-dependent l: -lactate dehydrogenase (ldha) and introduction ...201222228312
Construction of a novel sacB-based system for marker-free gene deletion in Corynebacterium glutamicum.Bacillus subtilis sacB gene with its 463bp upstream region including its native promoter has been used for marker-free gene deletion in Corynebacterium glutamicum, but the role of this upstream region is not clear. In this study, it was demonstrated that the upstream region of sacB failed to efficiently promote its expression in C. glutamicum, and the native promoter of sacB is weak in C. glutamicum. The expression level of sacB under its native promoter in C. glutamicum is not high enough for c ...201222100974
Efficient aerobic succinate production from glucose in minimal medium with Corynebacterium glutamicum.Corynebacterium glutamicum, an established industrial amino acid producer, has been genetically modified for efficient succinate production from the renewable carbon source glucose under fully aerobic conditions in minimal medium. The initial deletion of the succinate dehydrogenase genes (sdhCAB) led to an accumulation of 4.7 g l(-1) (40 mM) succinate as well as high amounts of acetate (125 mM) as by-product. By deleting genes for all known acetate-producing pathways (pta-ackA, pqo and cat) ace ...201222018023
phenol degradation activity and reusability of corynebacterium glutamicum coated with nh(2)-functionalized silica-encapsulated fe(3)o(4) nanoparticles.in this study, a novel method to immobilize and separate corynebacterium glutamicum for phenol degradation was developed using fe(3)o(4) nanoparticles (nps). the fe(3)o(4) nps were encapsulated with silica and functionalized with nh(2) groups to enhance their capacity to adsorb on the cell surface. the results showed that the nh(2)-functionalized silica-encapsulated fe(3)o(4) nps strongly adsorbed on the cell surface of c. glutamicum during 32d culture without any interruptions of their normal c ...201222093979
Physiology and global gene expression of a Corynebacterium glutamicum ?F(1)F(O)-ATP synthase mutant devoid of oxidative phosphorylation.A mutant of Corynebacterium glutamicum ATCC 13032 with a deletion of the atpBEFHAGDC genes encoding F(1)F(O)-ATP synthase was characterized. Whereas no growth was observed with acetate as sole carbon source, the ?F(1)F(O) mutant reached 47% of the growth rate and 65% of the biomass of the wild type during shake-flask cultivation in glucose minimal medium. Initially, the mutant strain showed a strongly increased glucose uptake rate accompanied by a high oxygen consumption rate and pyruvate secret ...201222050934
efflux permease cgacr3-1 of corynebacterium glutamicum is an arsenite-specific antiporter.resistance to arsenite (as(iii)) by cells is generally accomplished by arsenite efflux permeases from acr3 or arsb unrelated families. we analyzed the function of three acr3 proteins from corynebacterium glutamicum, cgacr3-1, cgacr3-2, and cgacr3-3. cgacr3-1 conferred the highest level of as(iii) resistance and accumulation in vivo. cgacr3-1 was also the most active when everted membranes vesicles from escherichia coli or c. glutamicum mutants were assayed for efflux with different energy source ...201222102279
postgenomic approaches to using corynebacteria as biocatalysts.corynebacterium glutamicum exhibits numerous ideal intrinsic attributes as a factory of primary and secondary metabolites. the versatile capabilities of this organism have long been implemented at the industrial scale to produce an array of amino acids at high yields and conversion rates, thereby enabling the development of an entire industry. the postgenomic era provides a new technological platform not only to further optimize the intrinsic attributes of c. glutamicum whole cells as biocatalys ...201222803796
corynebacterium glutamicum harbours a molybdenum cofactor-dependent formate dehydrogenase which alleviates growth inhibition in the presence of formate.here, we show that corynebacterium glutamicum atcc 13032 co-metabolizes formate when it is grown with glucose as the carbon and energy source. co(2) measurements during bioreactor cultivation and use of (13)c-labelled formate demonstrated that formate is almost completely oxidized to co(2). the deletion of fdhf (cg0618), annotated as formate dehydrogenase (fdh) and located in a cluster of genes conserved in the family corynebacteriaceae, prevented formate utilization. similarly, deletion of fdhd ...201222767548
effect of transport proteins on l-isoleucine production with the l-isoleucine-producing strain corynebacterium glutamicum yilw.previous studies have shown that the deletion of brnq from the corynebacterium glutamicum chromosome results in a significant reduction in l-isoleucine uptake rates, while overexpression of brnfe leads to enhanced l-isoleucine export rates. given that net excretion rates would be an important factor for high titers of l-isoleucine accumulation, we have tested the notion that decreased l-isoleucine uptake combined with increased l-isoleucine excretion will further improve high-yield strains that ...201222733295
overexpression of nad kinases improves the l-isoleucine biosynthesis in corynebacterium glutamicum ssp. lactofermentum.nadph is the key cofactor in l-isoleucine (ile) biosynthetic pathway. to increase the ile biosynthesis in corynebacterium glutamicum ssp. lactofermentum jhi3-156, nadph supply needs to be enhanced. here nad kinase, the key enzyme for the de novo biosynthesis of nadp(+) and nadph, were cloned and expressed in jhi3-156, and their influences on ile production were analysed. meanwhile, enzyme properties of nad kinase from jhi3-156 (cljppnk) were compared with that from c. glutamicum ssp. lactofermen ...201222664190
a gain-of-function mutation in gating of corynebacterium glutamicum ncgl1221 causes constitutive glutamate secretion.the a-to-v mutation at position 111 (a111v) in the mechanosensitive channel ncgl1221 (msccg) causes constitutive glutamate secretion in corynebacterium glutamicum. patch clamp experiments revealed that ncgl1221 (a111v) had a significantly smaller gating threshold than the wild-type counterpart and displayed strong hysteresis, suggesting that the gain-of-function mutation in the gating of ncgl1221 leads to the oversecretion of glutamate.201222610427
degradation and assimilation of aromatic compounds by corynebacterium glutamicum: another potential for applications for this bacterium?with the implementation of the well-established molecular tools and systems biology techniques, new knowledge on aromatic degradation and assimilation by corynebacterium glutamicum has been emerging. this review summarizes recent findings on degradation of aromatic compounds by c. glutamicum. among these findings, the mycothiol-dependent gentisate pathway was firstly discovered in c. glutamicum. other important knowledge derived from c. glutamicum would be the discovery of linkages among aromati ...201222588501
a tetracycline inducible expression vector for corynebacterium glutamicum allowing tightly regulable gene expression.here we report on the construction of a tetracycline inducible expression vector that allows a tightly regulable gene expression in corynebacterium glutamicum which is used in industry for production of small molecules such as amino acids. using the green fluorescent protein (gfp) as a reporter protein we show that this vector, named pclton1, is characterized by tight repression under non-induced conditions as compared to a conventional iptg inducible expression vector, and that it allows gradua ...201222587824
the development and application of a single-cell biosensor for the detection of l-methionine and branched-chain amino acids.the detection and quantification of specific metabolites in single bacterial cells is a major goal for industrial biotechnology. we have developed a biosensor based on the transcriptional regulator lrp that detects intracellular l-methionine and branched-chain amino acids in corynebacterium glutamicum. in assays, fluorescence output showed a linear relationship with cytoplasmic concentrations of the effector amino acids. in increasing order, the affinity of lrp for the amino acids is l-valine, l ...201222583745
negative role of wbla in response to oxidative stress in streptomyces coelicolor.in this study, we analyzed the oxidative stress response of wbla (whib-like gene a, sco3579), which was previously shown to be a global antibiotic down-regulator in streptomyces coelicolor. ever since a wbla ortholog named whca in corynebacterium glutamicum was found to play a negative role in the oxidative stress response, s. coelicolor wbla has been proposed to have a similar effect. a wbla-deletion mutant exhibited a less sensitive response to oxidative stress induced by diamide present in so ...201222573149
metabolic engineering and flux analysis of corynebacterium glutamicum for l-serine production.l-serine plays a critical role as a building block for cell growth, and thus it is difficult to achieve the direct fermentation of l-serine from glucose. in this study, corynebacterium glutamicum atcc 13032 was engineered de novo by blocking and attenuating the conversion of l-serine to pyruvate and glycine, releasing the feedback inhibition by l-serine to 3-phosphoglycerate dehydrogenase (pgdh), in combination with the co-expression of 3-phosphoglycerate kinase (pgk) and feedback-resistant pgdh ...201222566084
(l)-valine production with minimization of by-products' synthesis in corynebacterium glutamicum and brevibacterium flavum.corynebacterium glutamicum atcc13032 and brevibacterium flavum jv16 were engineered for l-valine production by over-expressing ilvebn ( r ) c genes at 31 °c in 72 h fermentation. different strategies were carried out to reduce the by-products' accumulation in l-valine fermentation and also to increase the availability of precursor for l-valine biosynthesis. the native promoter of ilva of c. glutamicum was replaced with a weak promoter mpilva (p-ilvam1cg) to reduce the biosynthetic rate of l-isol ...201222552525
purification and structure analysis of mycolic acids in corynebacterium glutamicum.corynebacterium glutamicum is widely used for producing amino acids. mycolic acids, the major components in the cell wall of c. glutamicum might be closely related to the secretion of amino acids. in this study, mycolic acids were extracted from 5 strains of c. glutamicum, including atcc 13032, atcc 13869, atcc 14067, l-isoleucine producing strain iwj-1, and l-valine producing strain vwj-1. structures of these mycolic acids were analyzed using thin layer chromatography and electrospray ionizatio ...201222538651
protein turnover quantification in a multilabeling approach: from data calculation to evaluation.liquid chromatography coupled to tandem mass spectrometry in combination with stable-isotope labeling is an established and widely spread method to measure gene expression on the protein level. however, it is often not considered that two opposing processes are responsible for the amount of a protein in a cell--the synthesis as well as the degradation. with this work, we provide an integrative, high-throughput method--from the experimental setup to the bioinformatics analysis--to measure synthes ...201222493176
quantification of proteome dynamics in corynebacterium glutamicum by (15)n-labeling and selected reaction monitoring.selected reaction monitoring allows quantitative measurements of proteins over several orders of magnitude in complex biological samples. here we present a targeted approach for quantification of 19 enzymes from corynebacterium glutamicum applying isotope dilution mass spectrometry coupled to high performance liquid chromatography (idms-lc-ms/ms). investigations of protein dynamics upon growth on acetate and glucose as sole carbon source shows highly stable peptide amounts for enzymes of the cen ...201222476105
global proteome survey of protocatechuate- and glucose-grown corynebacterium glutamicum reveals multiple physiological differences.corynebacterium glutamicum can utilize various monocyclic aromatic carbon sources, including protocatechuate, which is catabolized via the β-ketoadipate pathway. in order to obtain a global survey of occurring physiological adaptations on the proteome level, cytoplasmic and membrane fraction from cells grown on protocatechuate or glucose as sole carbon and energy source were compared. shotgun proteomics and relative protein quantification with metabolic isotope labeling and spectral counting wer ...201222450470
the role of corynebacterium glutamicum spia gene in whca-mediated oxidative stress gene regulation.the corynebacterium glutamicum whca protein, which inhibits the expression of oxidative stress response genes, is known to interact with the spia protein. in this study, we constructed and analyzed spia mutant cells with the goal of better understanding the function of the spia gene. a c. glutamicum strain overexpressing the spia gene showed retarded cell growth, which was caused by an increased sensitivity to oxidants. expression of the spia and whca genes was repressed by oxidant diamide, indi ...201222443283
improved l-lysine production with corynebacterium glutamicum and systemic insight into citrate synthase flux and activity.we here developed a series of corynebacterium glutamicum strains with gradual decreased specific citrate synthase (cs) activity and quantified in a multifaceted approach the consequences of residual activity on the transcriptome, metabolome, and fluxome level as well as on l-lysine formation and growth. we achieved an intended gradual l-lysine yield increase and recognized and overcame further new limitations in the l-lysine biosynthesis pathway to result in a strain with the highest yield repor ...201222392073
toward homosuccinate fermentation: metabolic engineering of corynebacterium glutamicum for anaerobic production of succinate from glucose and formate.previous studies have demonstrated the capability of corynebacterium glutamicum for anaerobic succinate production from glucose under nongrowing conditions. in this work, we have addressed two shortfalls of this process, the formation of significant amounts of by-products and the limitation of the yield by the redox balance. to eliminate acetate formation, a derivative of the type strain atcc 13032 (strain bol-1), which lacked all known pathways for acetate and lactate synthesis (δcat δpqo δpta- ...201222389371
gntr-type transcriptional regulator pckr negatively regulates the expression of phosphoenolpyruvate carboxykinase in corynebacterium glutamicum.the pck (cg3169) gene of corynebacterium glutamicum encodes a phosphoenolpyruvate carboxykinase (pepck). here, a candidate transcriptional regulator that binds to the promoter region of pck was detected using a dna affinity purification approach. an isolated protein was identified to be pckr (cg0196), a gntr family transcriptional regulator which consists of 253 amino acids with a mass of 27 kda as measured by peptide mass fingerprinting. the results of electrophoretic mobility shift assays veri ...201222366416
disruption of genes for the enhanced biosynthesis of α-ketoglutarate in corynebacterium glutamicum.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 aminotra ...201222356563
ndnr is an nad-responsive transcriptional repressor of the ndnr operon involved in nad de novo biosynthesis in corynebacterium glutamicum.the corynebacterium glutamicum ndnr gene, which is chromosomally located in a gene cluster involved in nad de novo biosynthesis, negatively regulates expression of the cluster genes, i.e. nada, nadc, nads and ndnr itself. although ndnr encodes a member of the recently identified nrtr family of transcriptional regulators, whether or not the ndnr protein directly regulates these nad biosynthesis genes remains to be verified. here, two ndnr binding sites in the promoter region of the ndnr-nada-nadc ...201222301909
characterization of the biotin uptake system encoded by the biotin-inducible bioymn operon of corynebacterium glutamicum.the amino acid-producing gram-positive corynebacterium glutamicum is auxotrophic for biotin although biotin ring assembly starting from the precursor pimeloyl-coa is still functional. it possesses accbc, the α-subunit of the acyl-carboxylases involved in fatty acid and mycolic acid synthesis, and pyruvate carboxylase as the only biotin-containing proteins. comparative genome analyses suggested that the putative transport system bioymn encoded by cg2147, cg2148 and cg2149 might be involved in bio ...201222243621
the gene encoding the alternative thymidylate synthase thyx is regulated by sigma factor sigb in corynebacterium glutamicum atcc 13032.both thya and thyx proteins catalyze the transfer of the methyl group from methylenetetrahydrofolate (ch(2) h(4) -folate) to dump, forming dtmp. to estimate the relative steady state expression levels of thya and thyx, western blot analysis was performed using thya or thyx antiserum on total protein from the wild-type, δthyx, and thyx-complemented strains of corynebacterium glutamicum. the level of thya decreased gradually during the stationary growth phase but that of thyx was maintained steadi ...201222224900
error propagation analysis for quantitative intracellular metabolomics.model-based analyses have become an integral part of modern metabolic engineering and systems biology in order to gain knowledge about complex and not directly observable cellular processes. for quantitative analyses, not only experimental data, but also measurement errors, play a crucial role. the total measurement error of any analytical protocol is the result of an accumulation of single errors introduced by several processing steps. here, we present a framework for the quantification of intr ...201224957773
metabolic engineering of the purine biosynthetic pathway in corynebacterium glutamicum results in increased intracellular pool sizes of imp and hypoxanthine.purine nucleotides exhibit various functions in cellular metabolism. besides serving as building blocks for nucleic acid synthesis, they participate in signaling pathways and energy metabolism. further, imp and gmp represent industrially relevant biotechnological products used as flavor enhancing additives in food industry. therefore, this work aimed towards the accumulation of imp applying targeted genetic engineering of corynebacterium glutamicum.201223092390
corynebacterium glutamicum csor acts as a transcriptional repressor of two copper/zinc-inducible p(1b)-type atpase operons.the mechanism of regulation of the expression of copa and copb, encoding putative copper-translocating p(1b)-type atpases in corynebacterium glutamicum, was investigated. the levels of copa and copb mrnas were upregulated in response to excess copper as well as excess zinc. disruption of csor, encoding a transcriptional regulator, resulted in constitutive expression of copa and copb. the csor protein bound to the promoter regions of the copa-csor and the cgr_0124-copb-cgr_0126 operon. in vitro d ...201223090582
sugar transport systems in corynebacterium glutamicum: features and applications to strain development.corynebacterium glutamicum uses the phosphoenolpyruvate-dependent sugar phosphotransferase system (pts) to take up and phosphorylate glucose, fructose, and sucrose, the major sugars from agricultural crops that are used as the primary feedstocks for industrial amino acid fermentation. this means that worldwide amino acid production using this organism has depended exclusively on the pts. recently, a better understanding not only of pts-mediated sugar uptake but also of global regulation associat ...201223081775
molecular mechanisms and metabolic engineering of glutamate overproduction in corynebacterium glutamicum.glutamate is a commercially important chemical. it is used as a flavor enhancer and is a major raw material for producing industrially useful chemicals. a coryneform bacterium, corynebacterium glutamicum, was isolated in 1956 by japanese researchers as a glutamate-overproducing bacterium and since then, remarkable progress in glutamate production has been made using this microorganism. currently, the global market for glutamate is over 2.5 million tons per year. glutamate overproduction by c. gl ...201223080255
analysis of corynebacterium glutamicum promoters and their applications.promoters are dna sequences which function as regulatory signals of transcription initiation catalyzed by rna polymerase. since promoters substantially influence levels of gene expression, they have become powerful tools in metabolic engineering. methods for their localization used in corynebacterium glutamicum and techniques for the analysis of their function are described in this review. c. glutamicum promoters can be classified according to the respective σ factors which direct rna polymerase ...201223080252
corynebacterium glutamicum zur acts as a zinc-sensing transcriptional repressor of both zinc-inducible and zinc-repressible genes involved in zinc homeostasis.zur is a zinc-dependent transcriptional repressor of zinc uptake systems in bacteria. in the present study, we examined the role of corynebacterium glutamicum zur in the zinc-inducible expression of two genes: one encoding a cation diffusion facilitator (zrf) and the other a metal-translocating p-type atpase (zra). both genes were shown to be involved in zinc resistance. disruption of the zur gene encoding zur resulted in constitutive expression of zrf and zra mrnas. an electrophoretic mobility ...201223061624
3' untranslated region-dependent degradation of the acea mrna, encoding the glyoxylate cycle enzyme isocitrate lyase, by rnase e/g in corynebacterium glutamicum.we previously reported that the corynebacterium glutamicum rnase e/g encoded by the rneg gene (ncgl2281) is required for the 5' maturation of 5s rrna. in the search for the intracellular target rnas of rnase e/g other than the 5s rrna precursor, we detected that the amount of isocitrate lyase, an enzyme of the glyoxylate cycle, increased in rneg knockout mutant cells grown on sodium acetate as the sole carbon source. rifampin chase experiments showed that the half-life of the acea mrna was about ...201223042181
the two-component system chrsa is crucial for haem tolerance and interferes with hrrsa in haem-dependent gene regulation in corynebacterium glutamicum.we recently showed that the two-component system (tcs) hrrsa plays a central role in the control of haem homeostasis in the gram-positive soil bacterium corynebacterium glutamicum. here, we characterized the function of another tcs of this organism, chrsa, which exhibits significant sequence similarity to hrrsa, and provide evidence for cross-regulation of the two systems. in this study, chrsa was shown to be crucial for haem resistance of c. glutamicum by activation of the putative haem-detoxif ...201223038807
the ppm operon is essential for acylation and glycosylation of lipoproteins in corynebacterium glutamicum.due to their contribution to bacterial virulence, lipoproteins and members of the lipoprotein biogenesis pathway represent potent drug targets. following translocation across the inner membrane, lipoprotein precursors are acylated by lipoprotein diacylglycerol transferase (lgt), cleaved off their signal peptides by lipoprotein signal peptidase (lsp) and, in gram-negative bacteria, further triacylated by lipoprotein n-acyl transferase (lnt). the existence of an active apolipoprotein n-acyltransfe ...201223029442
coordinated regulation of gnd, which encodes 6-phosphogluconate dehydrogenase, by the two transcriptional regulators gntr1 and rama in corynebacterium glutamicum.the transcriptional regulation of corynebacterium glutamicum gnd, encoding 6-phosphogluconate dehydrogenase, was investigated. two transcriptional regulators, gntr1 and rama, were isolated by affinity purification using gnd promoter dna. gntr1 was previously identified as a repressor of gluconate utilization genes, including gnd. involvement of rama in gnd expression had not been investigated to date. the level of gnd mrna was barely affected by the single deletion of rama. however, gnd expressi ...201223024346
implication of gluconate kinase activity in l-ornithine biosynthesis in corynebacterium glutamicum.with the purpose of generating a microbial strain for l-ornithine production in corynebacterium glutamicum, genes involved in the central carbon metabolism were inactivated so as to modulate the intracellular level of nadph, and to evaluate their effects on l-ornithine production in c. glutamicum. upon inactivation of the 6-phosphoglucoisomerase gene (pgi) in a c. glutamicum strain, the concomitant increase in intracellular nadph concentrations from 2.55 to 5.75 mmol g⁻¹ (dry cell weight) was ac ...201222987028
next-generation sequencing-based genome-wide mutation analysis of l-lysine-producing corynebacterium glutamicum atcc 21300 strain.in order to identify single nucleotide polymorphism and insertion/deletion mutations, we performed whole-genome re-sequencing of the enhanced l-lysine-producing corynebacterium glutamicum atcc 21300 strain. in total, 142 single nucleotide polymorphisms and 477 insertion/deletion mutations were identified in the atcc 21300 strain when compared to 3,434 predicted genes of the wild-type c. glutamicum atcc 13032 strain. among them, 110 transitions and 29 transversions of single nucleotide polymorphi ...201223124757
identification of a had superfamily phosphatase, hdpa, involved in 1,3-dihydroxyacetone production during sugar catabolism in corynebacterium glutamicum.corynebacterium glutamicum produces 1,3-dihydroxyacetone (dha) as metabolite of sugar catabolism but the responsible enzyme is yet to be identified. using a transposon mutant library, the gene hdpa (cgr_2128) was shown to encode a haloacid dehalogenase superfamily member that catalyzes dephosphorylation of dihydroxyacetone phosphate to produce dha. inactivation of hdpa led to a drastic decrease in dha production from each of glucose, fructose, and sucrose, indicating that hdpa is the main enzyme ...201223108048
analysis of in vivo function of predicted isoenzymes:a metabolomic approach.isoenzymes occur in all organisms. often, they are regulated with respect to environmental perturbations to assure metabolic flexibility. in bioinformatics-driven functional genome annotations, the presence of isoenzymes is frequently predicted. it is desirable to verify the functions of putative isoenzymes experimentally for a correct estimation of the metabolic capacities in a given organism. using metabolome analysis, we investigated two knockout mutants of putative shikimate dehydrogenases ( ...201223215805
[overexpression of corynebacterium glutamicum nad kinase improves l-isoleucine biosynthesis].nad kinase catalyzes the phosphorylation of coenzyme i [nad(h)] to form coenzyme ii [nadp(h)], and nadph is an important cofactor in l-isoleucine biosynthesis. in order to improve nadph supply, ppnk, the gene encoding nad kinase in corynebacterium glutamicum was cloned and separately expressed in an l-isoleucine synthetic strain, brevibacterium lactofermentum jhi3-156, by an inducible expression vector pdxw-8 and a constitutive expression vector pdxw-9. compared with the control strain jhi3-156/ ...201223289306
isolated microbial single cells and resulting micropopulations grow faster in controlled environments.singularized cells of pichia pastoris, hansenula polymorpha, and corynebacterium glutamicum displayed specific growth rates under chemically and physically constant conditions that were consistently higher than those obtained in populations. this highlights the importance of single-cell analyses by uncoupling physiology and the extracellular environment, which is now possible using the envirostat 2.0 concept.201222820335
transcriptional regulation of the operon encoding stress-responsive ecf sigma factor sigh and its anti-sigma factor rsha, and control of its regulatory network in corynebacterium glutamicum.the expression of genes in corynebacterium glutamicum, a gram-positive non-pathogenic bacterium used mainly for the industrial production of amino acids, is regulated by seven different sigma factors of rna polymerase, including the stress-responsive ecf-sigma factor sigh. the sigh gene is located in a gene cluster together with the rsha gene, putatively encoding an anti-sigma factor. the aim of this study was to analyze the transcriptional regulation of the sigh and rsha gene cluster and the ef ...201222943411
alternating-access mechanism in conformationally asymmetric trimers of the betaine transporter betp.betaine and na(+) symport has been extensively studied in the osmotically regulated transporter betp from corynebacterium glutamicum, a member of the betaine/choline/carnitine transporter family, which shares the conserved leut-like fold of two inverted structural repeats. betp adjusts its transport activity by sensing the cytoplasmic k(+) concentration as a measure for hyperosmotic stress via the osmosensing carboxy-terminal domain. betp needs to be in a trimeric state for communication between ...201222940865
sialic acid utilization by the soil bacterium corynebacterium glutamicum.the ability to use the sialic acid, n-acetylneuraminic acid, neu5ac, as a nutrient has been characterized in a number of bacteria, most of which are human pathogens that encounter this molecule because of its presence on mucosal surfaces. the soil bacterium corynebacterium glutamicum also has a full complement of genes for sialic acid catabolism, and we demonstrate that it can use neu5ac as a sole source of carbon and energy and isolate mutants with a much reduced growth lag on neu5ac. disruptio ...201222924979
quantitation of intracellular purine intermediates in different corynebacteria using electrospray lc-ms/ms.intermediates of the purine biosynthesis pathway play key roles in cellular metabolism including nucleic acid synthesis and signal mediation. in addition, they are also of major interest to the biotechnological industry as several intermediates either possess flavor-enhancing characteristics or are applied in medical therapy. in this study, we have developed an analytical method for quantitation of 12 intermediates from the purine biosynthesis pathway including important nucleotides and their co ...201222960872
molecular characterization of prpr, the transcriptional activator of propionate catabolism in corynebacterium glutamicum.the 2-methylcitrate cycle is used to metabolize propionate in corynebacterium glutamicum. the regulator, prpr (cg0800), of the prpdbc2 operon was identified and characterized. the regulator has no similarities to the up to now known prpr regulators from other organisms. growth of a δprpr mutant revealed severe growth deficits and a prolonged lag phase if propionate was present in the medium. transcriptome analyses demonstrated the inability of the δprpr strain to induce the prpdbc2 genes in the ...201121933687
The glgB-encoded glycogen branching enzyme is essential for glycogen accumulation in Corynebacterium glutamicum.Corynebacterium glutamicum transiently accumulates glycogen as carbon capacitor during the early exponential growth phase in media containing carbohydrates. In some bacteria glycogen is synthesized by the consecutive action of ADP-glucose pyrophosphorylase (GlgC), glycogen synthase (GlgA) and glycogen branching enzyme (GlgB). GlgC and GlgA of C. glutamicum have been shown to be necessary for glycogen accumulation in this organism. However, although cg1381 has been annotated as the putative C. gl ...201121903753
Site-Directed Mutagenesis Studies on the L: -Arginine-Binding Sites of Feedback Inhibition in N-Acetyl-L: -glutamate Kinase (NAGK) from Corynebacterium glutamicum.Arginine biosynthesis in Corynebacterium glutamicum proceeds via a pathway that is controlled by arginine through feedback inhibition of NAGK, the enzyme that converts N-acetyl-L: -glutamate (NAG) to N-acety-L: -glutamy-L: -phosphate. In this study, the gene argB encoding NAGK from C. glutamicum ATCC 13032 was site-directed, and the L: -arginine-binding sites of feedback inhibition in Cglu_NAGK are described. The N-helix and C-terminal residues were first deleted, and the results indicated that ...201122101454
Biotin protein ligase from Corynebacterium glutamicum: role for growth and L: -lysine production.Corynebacterium glutamicum is a biotin auxotrophic Gram-positive bacterium that is used for large-scale production of amino acids, especially of L: -glutamate and L: -lysine. It is known that biotin limitation triggers L: -glutamate production and that L: -lysine production can be increased by enhancing the activity of pyruvate carboxylase, one of two biotin-dependent proteins of C. glutamicum. The gene cg0814 (accession number YP_225000) has been annotated to code for putative biotin protein li ...201122159614
structural view of the regulatory subunit of aspartate kinase from mycobacterium tuberculosis.the aspartate kinase (ak) from mycobacterium tuberculosis (mtb) catalyzes the biosynthesis of aspartate family amino acids, including lysine, threonine, isoleucine and methionine. we determined the crystal structures of the regulatory subunit of aspartate kinase from mtb alone (referred to as mtbakβ) and in complex with threonine (referred to as mtbakβ-thr) at resolutions of 2.6 å and 2.0 å, respectively. mtbakβ is composed of two perpendicular non-equivalent act domains [aspartate kinase, chori ...201121976064
arabitol metabolism of corynebacterium glutamicum and its regulation by atlr.expression profiling of corynebacterium glutamicum in comparison to a derivative deficient of the transcriptional regulator atlr (previously known as sugr or mtlr) revealed eight genes showing more than fourfold higher mrna levels in the mutant. four of these are located in direct vicinity of the atlr gene, i.e., xylb, rbtt, mtld, and sixa, annotated as encoding xylulokinase, ribitol transporter, mannitol 2-dehydrogenase, and phosphohistidine phosphatase, respectively. transcriptional analysis i ...201122178972
bio-based production of chemicals, materials and fuels -corynebacterium glutamicum as versatile cell factory.since their discovery almost 60 years ago, corynebacterium glutamicum and related subspecies are writing a remarkable success story in industrial biotechnology. today, these gram-positive soil bacteria, traditionally well-known as excellent producers of l-amino acids are becoming flexible, efficient production platforms for various chemicals, materials and fuels. this development is intensively driven by systems metabolic engineering concepts integrating systems biology and synthetic biology int ...201122138494
Heterologous and homologous expression of the arginine biosynthetic argC~H cluster from Corynebacterium crenatum for improvement of L: -arginine production.The genes involved in L: -arginine biosynthesis in Corynebacterium crenatum are organized as the argCJBDFRGH cluster like in Corynebacterium glutamicum. However, the argC~H cluster of the C. crenatum SYPA 5-5, which is an industrialized L: -arginine producer, had a lethal mutation occurring in the ArgR repressor encoding gene. The argC~H cluster with an inactive argR was overexpressed in E. coli and C. crenatum. In the recombinant E. coli JM109 enzyme activities were increased, and more L: -argi ...201122009057
Impact of a new glucose utilization pathway in amino acid-producing Corynebacterium glutamicum.Corynebacterium glutamicum imports and phosphorylates glucose, fructose and sucrose by the phosphoenolpyruvate-dependent phosphotransferase carbohydrate uptake system (PTS). Recently, we have discovered how glucose can be utilized by C. glutamicum in a PTS-independent manner. PTS-independent glucose uptake is mediated by one of two inositol permeases (IolT1 or IolT2) and the second function of PTS, substrate phosphorylation, is catalyzed by one of two glucokinases (Glk or PpgK). PTS-deficient C. ...201122008639
conformational changes of the betaine transporter betp from corynebacterium glutamicum studied by pulse epr spectroscopy.the betaine transporter betp from corynebacterium glutamicum is activated by hyperosmotic stress critically depending on the presence and integrity of its sensory c-terminal domain. the conformational properties of the trimeric betp reconstituted in liposomes in the inactive state and during osmotic activation were investigated by site-directed spin labeling and electron paramagnetic resonance (epr) spectroscopy. comparison of intra- and intermolecular inter spin distance distributions obtained ...201122051018
quantitative lipid composition of cell envelopes of corynebacterium glutamicum elucidated through reverse micelle extraction.cells of the corynebacterium-nocardia-mycobacterium group of bacteria are surrounded by an outer membrane (om) containing mycolic acids that are covalently linked to the underlying arabinogalactan-peptidoglycan complex. this om presumably acts as a permeability barrier that imparts high levels of intrinsic drug resistance to some members of this group, such as mycobacterium tuberculosis, and its component lipids have been studied intensively in a qualitative manner over the years. however, the q ...201121876124
biochemical disclosure of the mycolate outer membrane of corynebacterium glutamicum.corynebacterineae is a specific sub-order of gram-positive bacteria that includes mycobacterium tuberculosis and corynebacterium glutamicum. their cell wall is composed of a heteropolymer of peptidoglycan (pg) linked to arabinogalactan (ag), which in turn, is covalently associated to an atypical outer membrane hereafter called mycomembrane (m). this latter structure has been visualized by cryoelectron microscopy of vitreous sections but its biochemical composition is still poorly defined, thereb ...201122123248
corynebacterium glutamicum as a potent biocatalyst for the bioconversion of pentose sugars to value-added products.corynebacterium glutamicum, the industrial microbe traditionally used for the production of amino acids, proved its value for the fermentative production of diverse products through genetic/metabolic engineering. a successful demonstration of the heterologous expression of arabinose and xylose utilization genes made them interesting biocatalysts for pentose fermentation, which are the main components in lignocellulosic hydrolysates. its ability to withstand substantial amount of general growth i ...201122094976
production of minicellulosomes for the enhanced hydrolysis of cellulosic substrates by recombinant corynebacterium glutamicum.although cellulosic materials of plant origin are the most abundant utilizable biomass resource, the amino acid-producing organism corynebacterium glutamicum can not utilize these materials. here we report the engineering of a c. glutamicum strain expressing functional minicellulosomes containing chimeric endoglucanase e bound to minicbpa from clostridium cellulovorans that can hydrolyze cellulosic materials. the chimeric endoglucanase e consists of the endoglucanase e catalytic backbone of clos ...201122112952
growth independent rhamnolipid production from glucose using the non-pathogenic pseudomonas putida kt2440.abstract:201121999513
diversity of metabolic shift in response to oxygen deprivation in corynebacterium glutamicum and its close relatives.oxygen-deprived corynebacterium glutamicum r cells remain metabolically active, producing considerable amounts of organic acids even when not actively growing. we compared the proficiencies of c. glutamicum and close relatives grown under aerobic conditions to metabolize glucose when deprived of oxygen. eight strains that readily consumed glucose without cell growth subsequently produced organic acids. among these, the glucose consumption rates of the two c. glutamicum strains (>40 mm/h) and cor ...201121327408
exploring the conformational dynamics and membrane interactions of porb from c. glutamicum: a multi-scale molecular dynamics simulation study.members of the gram-positive mycolata bacteria have unusual cell envelopes which help them to avoid the immune system and the effects of most antibiotics, whilst rendering them permeable to solutes of importance in industrial bioconversion. it is therefore of interest to understand the molecular mechanisms for this selective permeability. porb is an unusual porin from the outer membrane (om) of corynebacterium glutamicum. it has been proposed as an atypical a-helical, symmetrical homo-pentameric ...201121354102
efficient markerless gene replacement in corynebacterium glutamicum using a new temperature-sensitive plasmid.random chemical mutation of a corynebacterium glutamicum-escherichia coli shuttle vector derived from plasmid pcgr2 was done using hydroxylamine. it brought about amino acid substitutions g109d and e180k within the replicase superfamily domain of the plasmid's repa protein and rendered the plasmid highly unstable, especially at higher incubation temperatures. colony formation of c. glutamicum was consequently completely inhibited at 37°c but not at 25°c. g109 is a semi-conserved residue mutation ...201121362445
mixed glucose and lactate uptake by corynebacterium glutamicum through metabolic engineering.the corynebacterium glutamicum atcc 13032 lysc(fbr) strain was engineered to grow fast on racemic mixtures of lactate and to secrete lysine during growth on lactate as well as on mixtures of lactate and glucose. the wild-type c. glutamicum only grows well on l-lactate. overexpression of d-lactate dehydrogenase (dld) achieved by exchanging the native promoter of the dld gene for the stronger promoter of the sod gene encoding superoxide dismutase in c. glutamicum resulted in a duplication of bioma ...201121370474
mechanism of increased respiration in an h(+)-atpase-defective mutant of corynebacterium glutamicum.we previously reported that a spontaneous h(+)-atpase-defective mutant of corynebacterium glutamicum, f172-8, derived from c. glutamicum atcc 14067, showed enhanced glucose consumption and respiration rates. to investigate the genome-based mechanism of enhanced respiration rate in such c. glutamicum mutants, a-1, an h(+)-atpase-defective mutant derived from c. glutamicum atcc 13032, which harbors the same point mutation as f172-8, was used in this study. a-1 showed similar fermentation profiles ...201122188772
Structure of a highly NADP+-specific isocitrate dehydrogenase.Isocitrate dehydrogenase catalyzes the first oxidative and decarboxylation steps in the citric acid cycle. It also lies at a crucial bifurcation point between CO2-generating steps in the cycle and carbon-conserving steps in the glyoxylate bypass. Hence, the enzyme is a focus of regulation. The bacterial enzyme is typically dependent on the coenzyme nicotinamide adenine dinucleotide phosphate. The monomeric enzyme from Corynebacterium glutamicum is highly specific towards this coenzyme and the su ...201121931217
Enzymatic synthesis of S-adenosylhomocysteine: immobilization of recombinant S-adenosylhomocysteine hydrolase from Corynebacterium glutamicum (ATCC 13032).Recombinant S-adenosylhomocysteine hydrolase from Corynebacterium glutamicum (CgSAHase) was covalently bound to Eupergit® C. The maximum yield of bound protein was 91% and the catalytic efficiency was 96.9%. When the kinetic results for the immobilized enzyme were compared with those for the soluble enzyme, no decrease in the catalytic efficiency of the former was detected. Both soluble and immobilized enzymes showed similar optimum pH and temperature ranges. The reuse of immobilized CgSAHase ca ...201122202964
Purification, crystallization and preliminary X-ray diffraction studies of the arsenic repressor ArsR from Corynebacterium glutamicum.ArsR is a member of the SmtB/ArsR family of metalloregulatory proteins that regulate prokaryotic arsenic-resistance operons. Here, the crystallization and preliminary X-ray diffraction studies of a cysteine-free derivative of ArsR from Corynebacterium glutamicum (CgArsR-C15/16/55S) are reported. CgArsR-C15/16/55S was expressed, purified, crystallized and X-ray diffraction data were collected to 1.86 Å resolution. The protein crystallized in a tetragonal space group (P4), with unit-cell parameter ...201122139180
Corynebacterium glutamicum whcB, a stationary phase-specific regulatory gene.The function of whcB, one of the four whiB homologues of Corynebacterium glutamicum, was assessed. Cells carrying the P(180) -whcB clone, and thus overexpressing the whcB gene, showed retarded growth, probably due to increased sensitivity to oxidants, whereas cells lacking whcB (?whcB) did not. However, growth retardation was not observed in cells with additionally whcE deleted. Furthermore, the ?whcE phenotype, characterized by slow growth and sensitivity to oxidants, was reversed in cells carr ...201122098456
Engineered Corynebacterium glutamicum as an endotoxin-free platform strain for lactate-based polyester production.The first biosynthetic system for lactate (LA)-based polyesters was previously created in recombinant Escherichia coli (Taguchi et al. 2008). Here, we have begun efforts to upgrade the prototype polymer production system to a practical stage by using metabolically engineered Gram-positive bacterium Corynebacterium glutamicum as an endotoxin-free platform. We designed metabolic pathways in C. glutamicum to generate monomer substrates, lactyl-CoA (LA-CoA), and 3-hydroxybutyryl-CoA (3HB-CoA), for t ...201122127753
genome sequence of corynebacterium glutamicum s9114, a strain for industrial production of glutamate.here we report the genome sequence of corynebacterium glutamicum s9114, an industrial producer widely used in production of glutamate in china. preliminary comparison with the sequences of the corynebacterium glutamicum strains atcc 13032 and r revealed some notable mutagenesis that might be related to the high yield of glutamate.201121994927
Solution NMR structures reveal a distinct architecture and provide first structures for protein domain family PF04536.The protein family (Pfam) PF04536 is a broadly conserved domain family of unknown function (DUF477), with more than 1,350 members in prokaryotic and eukaryotic proteins. High-quality NMR structures of the N-terminal domain comprising residues 41-180 of the 684-residue protein CG2496 from Corynebacterium glutamicum and the N-terminal domain comprising residues 35-182 of the 435-residue protein PG0361 from Porphyromonas gingivalis both exhibit an a/ß fold comprised of a four-stranded ß-sheet, thre ...201122198206
Improving protein secretion of a transglutaminase-secreting Corynebacterium glutamicum recombinant strain on the basis of (13)C metabolic flux analysis.Corynebacterium glutamicum is known as a host species for amino acid production. This microorganism was recently noticed as a host that produces secreted proteins. In this study, we performed (13)C metabolic flux analysis ((13)C-MFA) on a recombinant C. glutamicum strain that secretes a heterologous transglutaminase (TGase) to improve TGase secretion. For the (13)C-MFA of a TGase-secreting C. glutamicum strain in batch cultivation, a (13)C-labeling experiment and measurement of mass isotopomer d ...201121903468
Corynebacterium glutamicum survives arsenic stress with arsenate reductases coupled to two distinct redox mechanisms.Arsenate reductases (ArsCs) evolved independently as a defence mechanism against toxic arsenate. In the genome of Corynebacterium glutamicum, there are two arsenic resistance operons (ars1 and ars2) and four potential genes coding for arsenate reductases (Cg_ArsC1, Cg_ArsC2, Cg_ArsC1' and Cg_ArsC4). Using knockout mutants, in vitro reconstitution of redox pathways, arsenic measurements and enzyme kinetics, we show that a single organism has two different classes of arsenate reductases. Cg_ArsC1 ...201122032722
deletion of the aconitase gene in corynebacterium glutamicum causes strong selection pressure for secondary mutations inactivating citrate synthase.the aconitase gene acn of corynebacterium glutamicum is regulated by four transcriptional regulators, indicating that the synthesis of this enzyme is carefully controlled. to understand the causes for this elaborate regulation, the properties of the δacn-1 deletion mutant were analyzed in detail. the mutant was glutamate auxotrophic in glucose minimal medium, showed a strong growth defect, and secreted large amounts of acetate. none of these phenotypes could be complemented by plasmid-encoded ac ...201121984793
current knowledge on isobutanol production with escherichia coli, bacillus subtilis and corynebacterium glutamicum.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 cha ...201122008938
improvement of the redox balance increases l-valine production by corynebacterium glutamicum under oxygen deprivation conditions.production of l-valine under oxygen deprivation conditions by corynebacterium glutamicum lacking the lactate dehydrogenase gene ldha and overexpressing the l-valine biosynthesis genes ilvbncde was repressed. this was attributed to imbalanced cofactor production and consumption in the overall l-valine synthesis pathway: two moles of nadh was generated and two moles of nadph was consumed per mole of l-valine produced from one mole of glucose. in order to solve this cofactor imbalance, the coenzyme ...201122138982
Phenotypic characterization of Corynebacterium glutamicum using elementary modes towards synthesis of amino acids.Elementary flux mode (EFM) analysis is a powerful tool to represent the metabolic network structure and can be further utilized for flux analysis. The method enables characterization and quantification of feasible phenotypes in microbes. EFM analysis was employed to characterize the phenotype of Corynebacterium glutamicum to yield various amino acids. The metabolic network of C. glutamicum yielded 62 elementary modes by incorporating the accumulation of amino acids namely, lysine, alanine, valin ...201122132055
genetic and biochemical characterization of corynebacterium glutamicum atp phosphoribosyltransferase and its three mutants resistant to feedback inhibition by histidine.atp phosphoribosyltransferase (atp-prt) catalyzes the condensation of atp and prpp at the first step of histidine biosynthesis and is regulated by a feedback inhibition from product histidine. here, we report the genetic and biochemical characterization of such an enzyme, hisg(cg), from corynebacterium glutamicum, including site-directed mutagenesis of the histidine-binding site for the first time. gene disruption and complementation experiments showed that hisg(cg) is essential for histidine bi ...201122172596
Ethambutol-mediated cell wall modification in recombinant Corynebacterium glutamicum increases the biotransformation rates of cyclohexanone derivatives.The effects of structural modification of cell wall on the biotransformation capability by recombinant Corynebacterium glutamicum cells, expressing the chnB gene encoding cyclohexanone monooxygenase of Acinetobacter calcoaceticus NCIMB 9871, were investigated. Baeyer-Villiger oxygenation of 2-(2'-acetoxyethyl) cyclohexanone (MW 170 Da) into R-7-(2'-acetoxyethyl)-2-oxepanone was used as a model reaction. The whole-cell biotransformation followed Michaelis-Menten kinetics. The V (max) and K (S) va ...201121909677
Negative transcriptional control of biotin metabolism genes by the TetR-type regulator BioQ in biotin-auxotrophic Corynebacterium glutamicum ATCC 13032.Genomic context analysis in actinobacteria revealed that biotin biosynthesis and transport (bio) genes are co-localized in several genomes with a gene encoding a transcription regulator of the TetR protein family, now named BioQ. Comparative analysis of the upstream regions of bio genes identified the common 13-bp palindromic motif TGAAC-N3-GTTAC as candidate BioQ-binding site. To verify the role of BioQ in controlling the transcription of bio genes, a deletion in the bioQ coding region (cg2309) ...201122178235
corynebacterium glutamicum as a host for synthesis and export of d-amino acids.a number of d-amino acids occur in nature, and there is growing interest in their function and metabolism, as well as in their production and use. here we use the well-established l-amino-acid-producing bacterium corynebacterium glutamicum to study whether d-amino acid synthesis is possible and whether mechanisms for the export of these amino acids exist. in contrast to escherichia coli, c. glutamicum tolerates d-amino acids added extracellularly. expression of argr (encoding the broad-substrate ...201121257776
positive transcriptional control of the pyridoxal phosphate biosynthesis genes pdxst by the mocr-type regulator pdxr of corynebacterium glutamicum atcc 13032.the pdxr (cg0897) gene of corynebacterium glutamicum atcc 13032 encodes a regulatory protein belonging to the mocr subfamily of gntr-type transcription regulators and consisting of an amino-terminal winged helix-turn-helix dna-binding domain and a carboxy-terminal aminotransferase-like domain. a defined deletion in the pdxr gene resulted in the decreased expression of the divergently orientated pdxst genes coding for the subunits of pyridoxal 5'-phosphate synthase. the pdxst mutant c. glutamicum ...201120847010
mutational analysis of splicing activities of ribonucleotide reductase α subunit protein from lytic bacteriophage p1201.a cp1201 rir1 intein is found in the ribonucleotide reductase alpha subunit (rnr α subunit) protein of lytic bacteriophage p1201 from corynebacterium glutamicum nchu 87078. this intein can be over-expressed and spliced in escherichia coli novablue cells. mutations of c539, the n-terminal residue of the c-extein in the cp1201 rir1 protein, led to the changes of pattern and level of protein-splicing activities. a g392s variant was found to be a temperature-sensitive protein with complete splicing ...201121210121
assessment of bacterial diversity in the cattle tick rhipicephalus (boophilus) microplus through tag-encoded pyrosequencing.ticks are regarded as the most relevant vectors of disease-causing pathogens in domestic and wild animals. the cattle tick, rhipicephalus (boophilus) microplus, hinders livestock production in tropical and subtropical parts of the world where it is endemic. tick microbiomes remain largely unexplored. the objective of this study was to explore the r. microplus microbiome by applying the bacterial 16s tag-encoded flx-titanium amplicon pyrosequencing (btefap) technique to characterize its bacterial ...201121211038
control of heme homeostasis in corynebacterium glutamicum by the two-component system hrrsa.the response regulator hrra of the hrrsa two-component system (previously named cgtsr11) was recently found to be repressed by the global iron-dependent regulator dtxr in corynebacterium glutamicum. here, we provide evidence that hrra mediates heme-dependent gene regulation in this nonpathogenic soil bacterium. growth experiments and dna microarray analysis revealed that c. glutamicum is able to use hemin as an alternative iron source and emphasize the involvement of the putative hemin abc trans ...201121217007
escherichia coli w as a new platform strain for the enhanced production of l-valine by systems metabolic engineering.a less frequently employed escherichia coli strain w, yet possessing useful metabolic characteristics such as less acetic acid production and high l-valine tolerance, was metabolically engineered for the production of l-valine. the ilva gene was deleted to make more pyruvate, a key precursor for l-valine, available for enhanced l-valine biosynthesis. the laci gene was deleted to allow constitutive expression of genes under the tac or trc promoter. the ilvbn(mut) genes encoding feedback-resistant ...201121191998
gene expression profiling of corynebacterium glutamicum during anaerobic nitrate respiration: induction of the sos response for cell survival.the gene expression profile of corynebacterium glutamicum under anaerobic nitrate respiration revealed marked differences in the expression levels of a number of genes involved in a variety of cellular functions, including carbon metabolism and respiratory electron transport chain, compared to the profile under aerobic conditions using dna microarrays. many sos genes were upregulated by the shift from aerobic to anaerobic nitrate respiration. an elongated cell morphology, similar to that induced ...201121239583
from zero to hero--design-based systems metabolic engineering of corynebacterium glutamicum for l-lysine production.here, we describe the development of a genetically defined strain of l-lysine hyperproducing corynebacterium glutamicum by systems metabolic engineering of the wild type. implementation of only 12 defined genome-based changes in genes encoding central metabolic enzymes redirected major carbon fluxes as desired towards the optimal pathway usage predicted by in silico modeling. the final engineered c. glutamicum strain was able to produce lysine with a high yield of 0.55 g per gram of glucose, a t ...201121241816
sigma factors and promoters in corynebacterium glutamicum.the corynebacterium glutamicum genome codes for 7 sigma subunits (factors) of rna polymerase (rnap): primary sigma factor siga (s(a)), primary-like sigb and 5 other alternative sigma factors (sigc, sigd, sige, sigh and sigm). each sigma factor is responsible for recognizing promoters of genes belonging to a regulon (sigmulon) involved in specific functions of the cell. most promoters of c. glutamicum housekeeping genes are recognized by rnap+s(a), whereas s(b) is involved in transcription of a l ...201121277915
transcriptional regulators of multiple genes involved in carbon metabolism in corynebacterium glutamicum.corynebacterium glutamicum, a high-gc gram-positive soil bacterium, has been used in development of bioprocesses for production of various compounds such as amino acids, organic acids, and alcohols. recently, several transcriptional regulators, each of which is involved in multiple carbon metabolic pathways in this bacterium, have been identified and characterized. these regulators appear to form a complicated network mediating coordinated expression of a number of metabolic genes for efficient ...201121277916
structural asymmetry in a trimeric na+/betaine symporter, betp, from corynebacterium glutamicum.the na(+)-coupled symporter betp catalyzes the uptake of the compatible solute betaine in the soil bacterium corynebacterium glutamicum. betp also senses hyperosmotic stress and regulates its own activity in response to stress level. we determined a three-dimensional (3d) map (at 8 å in-plane resolution) of a constitutively active mutant of betp in a c. glutamicum membrane environment by electron cryomicroscopy of two-dimensional crystals. the map shows that the constitutively active mutant, whi ...201121281647
impact of improved potassium accumulation on ph homeostasis, membrane potential adjustment and survival of corynebacterium glutamicum.metal ion uptake is crucial for all living cells and an essential part of cellular bioenergetic homeostasis. in this study the uptake and the impact of the most abundant internal cation, potassium, were investigated in actinobacteria, a group of high g+c gram-positives with a number of prominent biotechnologically and medically important members. genome analyses revealed a variety of different potassium uptake systems in this monophyletic group ranging from potassium channels common in virtually ...201121295539
metabolic engineering of corynebacterium glutamicum for production of 1,5-diaminopentane from hemicellulose.in the present work, the bio-based production of 1,5-diaminopentane (cadaverine), an important building block for bio-polyamides, was extended to hemicellulose a non-food raw material. for this purpose, the metabolism of 1,5-diaminopentane-producing corynebacterium glutamicum was engineered to the use of the c(5) sugar xylose. this was realized by heterologous expression of the xyla and xylb genes from escherichia coli, mediating the conversion of xylose into xylulose 5-phosphate (an intermediat ...201121298810
glutamate production from ß-glucan using endoglucanase-secreting corynebacterium glutamicum.we demonstrate glutamate production from ß-glucan using endoglucanase (eg)-expressing corynebacterium glutamicum. the signal sequence tora derived from escherichia coli k12, which belongs to the tat pathway, was suitable for secreting eg of clostridium thermocellum using c. glutamicum as a host. using the tora signal sequence, endoglucanase from clostridium cellulovorans 743b was successfully expressed, and the secreted eg produced 123 mg of reducing sugar from 5 g of ß-glucan at 30 °c for 72 h, ...201121305281
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