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a high-resolution titrator: a new approach to studying binding sites of microbial biosorbents.the high-resolution potentiometric titration was used as a physico-chemical method to study the acid properties of selected biosorbent materials in order to quantify the functional acidic groups for sorption and to determine their affinities by considering their partial or total ionization equilibrium reactions. the gran's method and the henderson-hasselbach's equation were employed in establishing the partition of the total acidity as associated with strong, weak and very weak acidic chemical a ...200415707630
purification of l-lysine in simulated moving bed and fixed-bed chromatography.l-lysine was produced by a microbial process utilizing a corynebacterium glutamicum (atcc 21799) strain. l-lysine was purified from the cultivated medium by fixed-bed and simulated moving bed (smb) chromatography. the separation conditions including ph, eluent concentration and lys+ and lys2+ adsorption isotherms were studied in batch adsorption. the column capacity, eluent flow rate and eluent concentration have been studied in fixed-bed chromatography. maximum purification rate of lysine was o ...200415709427
mapping the membrane proteome of corynebacterium glutamicum.in order to avoid the specific problems with intrinsic membrane proteins in proteome analysis, a new procedure was developed which is superior to the classical two-dimensional polyacrylamide gel electrophoresis (2-d page) method in terms of intrinsic membrane proteins. for analysis of the membrane proteome from corynebacterium glutamicum, we replaced the first separation dimension, i.e., the isoelectric focusing step, by anion-exchange chromatography, followed by sodium dodecyl sulfate (sds)-pag ...200515717325
analyses of enzyme ii gene mutants for sugar transport and heterologous expression of fructokinase gene in corynebacterium glutamicum atcc 13032.corynebacterium glutamicum atcc 13032 has four enzyme ii (eii) genes of the phosphotransferase system in its genome encoding transporters for sucrose, glucose, fructose, and an unidentified eii. to analyze the function of these eii genes, they were inactivated via homologous recombination and the resulting mutants characterized for sugar utilization. whereas the sucrose eii was the only transport system for sucrose in c. glutamicum, fructose and glucose were each transported by a second transpor ...200515766777
gene expression systems in corynebacteria.corynebacterium belongs to a group of gram-positive bacteria having moderate to high g+c content, the other members being mycobacterium, nocardia, and rhodococcus. considerable information is now available on the plasmids, gene regulatory elements, and gene expression in corynebacteria, especially in soil corynebacteria such as corynebacterium glutamicum. these bacteria are non-pathogenic and, unlike bacillus and streptomyces, are low in proteolytic activity and thus have the potential of becomi ...200515766862
efficient production of (2s)-flavanones by escherichia coli containing an artificial biosynthetic gene cluster.for the fermentative production of plant-specific flavanones (naringenin, pinocembrin) by escherichia coli, a plasmid was constructed which carried an artificial biosynthetic gene cluster, including pal encoding a phenylalanine ammonia-lyase from a yeast, scccl encoding a cinnamate/coumarate:coa ligase from the actinomycete streptomyces coelicolor a3(2), chs encoding a chalcone synthase from a licorice plant and chi encoding a chalcone isomerase from the pueraria plant. the recombinant e. coli c ...200515770480
comparative study of flux redistribution of metabolic pathway in glutamate production by two coryneform bacteria.in amino acid production by coryneform bacteria, study on relationship between change in enzyme activities and production of a target amino acid is important. in glutamate production, kawahara et al. discovered that the effect of decrease in 2-oxoglutamate dehydrogenase complex (odhc) on glutamate production is essential (kawahara et al., biosci. biotechnol. biochem. 61(7) (1997) 1109). significant reduction of the odhc activity was observed in the cells under the several glutamate-productive co ...200515781416
overexpression of the ftsz gene from corynebacterium glutamicum (brevibacterium lactofermentum) in escherichia coli.our goal in this work was to overexpress the essential cell division ftsz protein from corynebacterium glutamicum (brevibacterium lactofermentum) (ftszcg) in escherichia coli to produce anti-ftszcg polyclonal antibodies. previous results from our laboratory showed that ftszcg was not expressed in e. coli in a sufficient amount to purify ftszcg. however, when ftszcg (without upstream sequences) was transcriptionally fused to the t7 promoter, different truncated ftszcg proteins (28-32 kda) were ov ...200515782238
in vivo quantification of intracellular amino acids and intermediates of the methionine pathway in corynebacterium glutamicum. 200515802143
characterization of ltsa from rhodococcus erythropolis, an enzyme with glutamine amidotransferase activity.the nocardioform actinomycete rhodococcus erythropolis has a characteristic cell wall structure. the cell wall is composed of arabinogalactan and mycolic acid and is highly resistant to the cell wall-lytic activity of lysozyme (muramidase). in order to improve the isolation of recombinant proteins from r. erythropolis host cells (n. nakashima and t. tamura, biotechnol. bioeng. 86:136-148, 2004), we isolated two mutants, l-65 and l-88, which are susceptible to lysozyme treatment. the lysozyme sen ...200515805504
theoretical aspects of 13c metabolic flux analysis with sole quantification of carbon dioxide labeling.the potential of using sole respirometric co2 labeling measurement for 13c metabolic flux analysis was investigated by metabolic simulations. for this purpose a model was created, considering all co2 forming and consuming reactions in the central catabolic and anabolic pathways. to facilitate the interpretation of the simulation results, the underlying metabolic network was parameterized by physiologically meaningful flux parameters such as flux partitioning ratios at metabolic branch points and ...200515833440
cre/loxp-mediated deletion system for large genome rearrangements in corynebacterium glutamicum.genome rearrangement is an increasingly important technique to facilitate the understanding of genome functions. a cre/loxp-mediated deletion system for large-scale genome rearrangements in corynebacterium glutamicum was developed. by comparative analysis of c. glutamicum r and c. glutamicum 13032 genomes, distinct 14.5-kb and 56-kb regions not essential for cell survival were identified and targeted for deletion. by homologous recombination, loxp sites were integrated at each end of the target ...200415834716
multiple large segment deletion method for corynebacterium glutamicum.a precise and scarless genome excision method, employing the cre/loxp system in concert with double-strand break (dsb)-stimulated intramolecular recombination was developed. the dsbs were mediated by the restriction endonuclease, i-scei. it permitted multiple deletions of independent 14-, 43-, and 10-kb-long genomic regions on the corynebacterium glutamicum genome. accuracy of deletion was confirmed by the loss of marker genes, pcr, and sequencing of new genome joints. eleven, 58, and 4 genes we ...200515843930
the mcbr repressor modulated by the effector substance s-adenosylhomocysteine controls directly the transcription of a regulon involved in sulphur metabolism of corynebacterium glutamicum atcc 13032.in a recent proteomics study we have shown that the mcbr gene of corynebacterium glutamicum atcc 13032 most probably encodes a transcriptional repressor of the tetr type, which regulates the expression of at least six genes involved in the synthesis of sulphur-containing amino acids. by means of dna microarray hybridizations we detected 86 genes with enhanced transcription in an mcbr mutant when compared with the wild-type strain. bioinformatic analysis identified the inverted repeat 5'-tagac-n6 ...200515853877
adaptation of corynebacterium glutamicum to ammonium limitation: a global analysis using transcriptome and proteome techniques.theresponse of corynebacterium glutamicum to ammonium limitation was studied by transcriptional and proteome profiling of cells grown in a chemostat. our results show that ammonium-limited growth of c. glutamicum results in a rearrangement of the cellular transport capacity, changes in metabolic pathways for nitrogen assimilation, amino acid biosynthesis, and carbon metabolism, as well as a decreased cell division. since transcription at different growth rates was studied, it was possible to dis ...200515870326
ethambutol, a cell wall inhibitor of mycobacterium tuberculosis, elicits l-glutamate efflux of corynebacterium glutamicum.corynebacterium glutamicum is used for the large-scale production of l-glutamate, but the efflux of this amino acid is poorly understood. this study shows that addition of ethambutol (emb) to growing cultures of c. glutamicum causes l-glutamate efflux at rates of up to 15 nmol min(-1) (mg dry wt)(-1), whereas in the absence of emb, no efflux occurs. emb is used for the treatment of mycobacterium tuberculosis, and at a molecular level it targets a series of arabinosyltransferases (embcab). the si ...200515870446
functional analysis of sigh expression in corynebacterium glutamicum.the sigh gene of corynebacterium glutamicum encodes ecf sigma factor sigmah. the gene apparently plays an important role in other stress responses as well as heat stress response. in this study, we found that deleting the sigh gene made c. glutamicum cells sensitive to the thiol-specific oxidant diamide. in the sigh mutant strain, the activity of thioredoxin reductase markedly decreased, suggesting that the trxb gene encoding thioredoxin reductase is probably under the control of sigmah. the exp ...200515883048
purification and characterization of succinate:menaquinone oxidoreductase from corynebacterium glutamicum.succinate:menaquinone oxidoreductase from corynebacterium glutamicum, a high-g+c, gram-positive bacterium, was purified to homogeneity. the enzyme contained two heme b molecules and three polypeptides with apparent molecular masses of 67, 29 and 23 kda, which corresponded to sdha (flavoprotein), sdhb (iron-sulfur protein), and sdhc (membrane anchor protein), respectively. in non-denaturating polyacrylamide gel electrophoresis, the enzyme migrated as a single band with an apparent molecular mass ...200515883782
characterization of methionine export in corynebacterium glutamicum.corynebacterium glutamicum is known for its effective excretion of amino acids under particular metabolic conditions. concomitant activities of uptake and excretion systems would create an energy-wasting futile cycle; amino acid export systems are therefore tightly regulated. we have used a dna microarray approach to identify genes for membrane proteins which are overexpressed under conditions of elevated cytoplasmic concentrations of methionine. one of these genes was brnf, coding for the large ...200515901702
the crucial role of trehalose and structurally related oligosaccharides in the biosynthesis and transfer of mycolic acids in corynebacterineae.trehalose (alpha-d-glucopyranosyl-alpha'-d-glucopyranoside) is essential for the growth of the human pathogen mycobacterium tuberculosis but not for the viability of the phylogenetically related corynebacteria. to determine the role of trehalose in the physiology of these bacteria, the so-called corynebacterineae, mutant strains of corynebacterium glutamicum unable to synthesize trehalose due to the knock-out of the genes of the three pathways of trehalose biosynthesis, were biochemically analyz ...200515901732
corynebacterium glutamicum glyceraldehyde-3-phosphate dehydrogenase isoforms with opposite, atp-dependent regulation.corynebacterium glutamicum gapa and gapb encode glyceraldehyde-3-phosphate dehydrogenases (gapdhs) that differ in molecular weight and activity in the presence of atp. comparative genome analysis revealed that gapa, the product of gapa, represented the canonical gapdh that is highly conserved across the three major life forms. gapb, with an additional 110-residue-long sequence upstream of its gapdh-specific domain, was homologous only to select microbial putative gapdhs. upon gene disruption, th ...200415925900
rational design of a corynebacterium glutamicum pantothenate production strain and its characterization by metabolic flux analysis and genome-wide transcriptional profiling.a "second-generation" production strain was derived from a corynebacterium glutamicum pantothenate producer by rational design to assess its potential to synthesize and accumulate the vitamin pantothenate by batch cultivation. the new pantothenate production strain carries a deletion of the ilva gene to abolish isoleucine synthesis, the promoter down-mutation p-ilvem3 to attenuate ilve gene expression and thereby increase ketoisovalerate availability, and two compatible plasmids to overexpress t ...200515933028
large-scale engineering of the corynebacterium glutamicum genome.the engineering of corynebacterium glutamicum is important for enhanced production of biochemicals. to construct an improved c. glutamicum genome, we developed a precise genome excision method based on the cre/loxp recombination system and successfully deleted 11 distinct genomic regions identified by comparative analysis of c. glutamicum genomes. despite the loss of several predicted open reading frames, the mutant cells exhibited normal growth under standard laboratory conditions. with a total ...200515933044
dna microarray analysis of the nitrogen starvation response of corynebacterium glutamicum.nitrogen is an essential component of nearly all of the complex macromolecules in a bacterial cell, e.g. proteins, nucleic acids, and cell wall components. accordingly, most prokaryotes have developed elaborate control mechanisms to provide an optimal supply of nitrogen for cellular metabolism and to cope with situations of nitrogen limitation. in this communication, a global analysis of the corynebacterium glutamicum nitrogen starvation response by transcriptional profiling using dna microarray ...200515935503
the individual and common repertoire of dna-binding transcriptional regulators of corynebacterium glutamicum, corynebacterium efficiens, corynebacterium diphtheriae and corynebacterium jeikeium deduced from the complete genome sequences.the genus corynebacterium includes gram-positive microorganisms of great biotechnologically importance, such as corynebacterium glutamicum and corynebacterium efficiens, as well as serious human pathogens, such as corynebacterium diphtheriae and corynebacterium jeikeium. although genome sequences of the respective species have been determined recently, the knowledge about the repertoire of transcriptional regulators and the architecture of global regulatory networks is scarce. here, we apply a c ...200515938759
heterologous expression of escherichia coli ppsa (phosphoenolpyruvate synthetase) and galu (udp-glucose pyrophosphorylase) genes in corynebacterium glutamicum, and its impact on trehalose synthesis.trehalose is a disaccharide with a wide range of applications in the food industry. we recently proposed a strategy for trehalose production based on a corynebacterium glutamicum strain expressing the escherichia coli enzyme udp-glucose pyrophosphorylase (galu). biochemical network analysis suggest a further bottleneck for trehalose synthesis resulting from the coupling of phosphotransferase (pts) mediated glucose uptake, and glucose catabolism in c. glutamicum. to overcome this coupling, we pro ...200515949962
strain improvement by metabolic engineering: lysine production as a case study for systems biology.a central goal of systems biology is the elucidation of cell function and physiology through the integrated use of broad based genomic and physiological data. such systemic approaches have been employed extensively in the past, as they are a central element of metabolic flux analysis, the distribution of kinetic control in pathways, and the key differentiating characteristic of metabolic engineering. in one case study, these tools have been applied to the improvement of lysine-producing strains ...200515961038
complete genome sequence and analysis of the multiresistant nosocomial pathogen corynebacterium jeikeium k411, a lipid-requiring bacterium of the human skin flora.corynebacterium jeikeium is a "lipophilic" and multidrug-resistant bacterial species of the human skin flora that has been recognized with increasing frequency as a serious nosocomial pathogen. here we report the genome sequence of the clinical isolate c. jeikeium k411, which was initially recovered from the axilla of a bone marrow transplant patient. the genome of c. jeikeium k411 consists of a circular chromosome of 2,462,499 bp and the 14,323-bp bacteriocin-producing plasmid pkw4. the chromos ...200515968079
vanillate metabolism in corynebacterium glutamicum.corynebacterium glutamicum, a gram-positive soil bacterium belonging to the mycolic acids-containing actinomycetes, is able to use the lignin degradation products ferulate, vanillate, and protocatechuate as sole carbon sources. the gene cluster responsible for vanillate catabolism was identified and characterized. the vanab genes encoding vanillate demethylase are organized in an operon together with the vank gene, coding for a transport system most likely responsible for protocatechuate uptake. ...200515971090
[purification and characterization of glutamate dehydrogenase. from corynebacterium glutamicum s9114].glutamate dehydrogenase (gdh) is a key enzyme in the biosynthesis of glutamate. the gdhs from corynebacterium glutamicum s9114 the most commonly used strain in glutamate fermentation, were purified and their molecular structures and properties characterized. the coenzymes were also studied in the hope to increase glutamate production. cells were harvested at mid-exponential phase by centrifugation and washed with tris-hcl buffer containing dtt and edta (ph 7.5). the cells were then disrupted usi ...200315971587
[metabolic flux analysis of l-valine fermentation in corynebacterium glutamicum].in industrial fermentation of amino acids the cells are often forced to synthesize the biochemicals excessive of their physiological needs. the knowledge of metabolic networks and their regulation relevant usually come from biochemical research, especially from enzymology, not from engineering study. to enrich the knowledge of metabolic sub-network of l-valine syntheses for higher production of l-valine, corynebacterium glutamicum as1.495 and its genetic derivatives aa361, aat231, aatv341 were u ...200415971614
the transcriptional activator clgr controls transcription of genes involved in proteolysis and dna repair in corynebacterium glutamicum.expression of the structural genes encoding the atp-dependent proteases clpcp and lon in corynebacterium glutamicum and streptomyces lividans is activated by the transcriptional regulator clgr in response to yet unknown environmental stimuli. as it was not known whether clgr controls expression of additional genes we used dna microarrays in order to comprehensively define the clgr regulon in c. glutamicum. the mrna levels of 16 genes decreased >/= 2-fold in a deltaclgrdeltaclpc mutant (clgr abse ...200515978086
high expression with corynebacterium glutamicum for nuclear magnetic resonance sample preparation. 200515979559
lysine and glutamate production by corynebacterium glutamicum on glucose, fructose and sucrose: roles of malic enzyme and fructose-1,6-bisphosphatase.in the biotechnological production of l-lysine and l-glutamate by corynebacterium glutamicum media based on glucose, fructose or sucrose are typically used. glutamate production by c. glutamicum was very similar on glucose, fructose, glucose plus fructose and sucrose. in contrast, lysine production of genetically defined c. glutamicum strains was significantly higher on glucose than on the other carbon sources. to test whether malic enzyme or fructose-1,6-bisphosphatase might limit growth and ly ...200515979917
application of model discriminating experimental design for modeling and development of a fermentative fed-batch l-valine production process.a model discriminating experimental design approach for fed-batch processes has been developed and applied to the fermentative production of l-valine by a genetically modified corynebacterium glutamicum strain possessing multiple auxotrophies as an example. being faced with the typical situation of uncertain model information based on preliminary experiments, model discriminating design was successfully applied to improve discrimination between five competing models. within the same modeling and ...200515984033
the amrg1 gene is involved in the activation of acetate in corynebacterium glutamicum.during growth of corynebacterium glutamicum on acetate as its carbon and energy source, the expression of the pta-ack operon is induced, coding for the acetate-activating enzymes, which are phosphotransacetylase (pta) and acetate kinase (ak). by transposon rescue, we identified the two genes amrg1 and amrg2 found in the deregulated transposon mutant c. glutamicum g25. the amrg1 gene (ncbi-accession: af532964) has a size of 732 bp, encoding a polypeptide of 243 amino acids and apparently is parti ...200515986882
chill activation of compatible solute transporters in corynebacterium glutamicum at the level of transport activity.the gram-positive soil bacterium corynebacterium glutamicum harbors four osmoregulated secondary uptake systems for compatible solutes, betp, ectp, lcop, and prop. when reconstituted in proteoliposomes, betp was shown to sense hyperosmotic conditions via the increase in luminal k(+) and to respond by instant activation. to study further putative ways of stimulus perception and signal transduction, we have investigated the responses of ectp, lcop, and betp, all belonging to the betaine-carnitine- ...200515995189
two functional fas-i type fatty acid synthases in corynebacterium glutamicum.the lipid-rich corynebacterianeae, to which corynebacterium glutamicum and mycobacterium species belong, produce both fatty acids and mycolic acids. compared with most other bacteria, c. glutamicum possesses two fatty acid synthases, encoded by fasa (8907 kb; fas-ia) and fasb (8988 kb; fas-ib). here, it was shown by mutational analyses that fasa is essential but fasb is not. however, in a fasa background, the fasb mutation results in a slightly reduced growth yield, l-glutamate production is inc ...200516000732
porh, a new channel-forming protein present in the cell wall of corynebacterium efficiens and corynebacterium callunae.corynebacterium callunae and corynebacterium efficiens are close relatives of the glutamate-producing mycolata species corynebacterium glutamicum. the properties of the pore-forming proteins, extracted by organic solvents, were studied. the cell extracts contained channel-forming proteins that formed ion-permeable channels with a single-channel conductance of about 2 to 3 ns in 1 m kcl in a lipid bilayer assay. the corresponding proteins from both corynebacteria were purified to homogeneity and ...200516000733
functional identification of novel genes involved in the glutathione-independent gentisate pathway in corynebacterium glutamicum.corynebacterium glutamicum used gentisate and 3-hydroxybenzoate as its sole carbon and energy source for growth. by genome-wide data mining, a gene cluster designated ncg12918-ncg12923 was proposed to encode putative proteins involved in gentisate/3-hydroxybenzoate pathway. genes encoding gentisate 1,2-dioxygenase (ncg12920) and fumarylpyruvate hydrolase (ncg12919) were identified by cloning and expression of each gene in escherichia coli. the gene of ncg12918 encoding a hypothetical protein (nc ...200516000747
[glyoxylate cycle is required for the overproduction of glutamate but is not essential for corynebacterium glutamicum growth on glucose].the glyoxylate cycle was hypothesed to be indispensable for glutamate overproduction in coryneform bacteria, for it was thought to fulfill anaplerotic functions and to supply energy during the growth phase. during glutamate overproduction phase, however, it has been noted that the high level of the cycle is detrimental to the glutamate production. in order to clarify the relationship between the glutamate production and the glyoxylate cycle, a chromosomal acea-disrupted mutant of wild-type c. gl ...200516013488
deletion of cg-emb in corynebacterianeae leads to a novel truncated cell wall arabinogalactan, whereas inactivation of cg-ubia results in an arabinan-deficient mutant with a cell wall galactan core.the cell wall of mycobacterium tuberculosis has a complex ultrastructure that consists of mycolic acids connected to peptidoglycan via arabinogalactan (ag) and abbreviated as the magp complex. the magp complex is crucial for the survival and pathogenicity of m. tuberculosis and is the target of several anti-tubercular agents. apart from sharing a similar magp and the availability of the complete genome sequence, corynebacterium glutamicum has proven useful in the study of orthologous m. tubercul ...200516040600
the impact of fluid mechanical stress on corynebacterium glutamicum during continuous cultivation in an agitated bioreactor.the effect of mechanical stresses generated by an extreme agitation intensity or a high aeration rate on growth parameters and cell physiology were studied during continuous cultivation of the gram-positive bacterium corynebacterium glutamicum. it is concluded that variations in agitation, aeration rate, or do2 concentrations down to about 1% of saturation do not damage the bacterial cells or cause a significant change in physiological response, as measured by flow cytometry, even though the cel ...200516049736
altered morphology produced by ftsz expression in corynebacterium glutamicum atcc 13869.corynebacterium glutamicum is a gram-positive bacterium that lacks the cell division ftsa protein and actin-like mreb proteins responsible for determining cylindrical cell shape. when the cell division ftsz gene from c. glutamicum (ftsz(cg)) was cloned in different multicopy plasmids, the resulting constructions could not be introduced into c. glutamicum; it was assumed that elevated levels of ftsz(cg) result in lethality. the presence of a truncated ftsz(cg) and a complete ftsz(cg) under the co ...200516079335
phosphate starvation-inducible gene usha encodes a 5' nucleotidase required for growth of corynebacterium glutamicum on media with nucleotides as the phosphorus source.phosphorus is an essential component of macromolecules, like dna, and central metabolic intermediates, such as sugar phosphates, and bacteria possess enzymes and control mechanisms that provide an optimal supply of phosphorus from the environment. udp-sugar hydrolases and 5' nucleotidases may play roles in signal transduction, as they do in mammals, in nucleotide salvage, as demonstrated for usha of escherichia coli, or in phosphorus metabolism. the corynebacterium glutamicum gene usha was found ...200516085822
key enzymes of the protocatechuate branch of the beta-ketoadipate pathway for aromatic degradation in corynebacterium glutamicum.although the protocatechuate branch of the beta-ketoadipate pathway in gram+ bacteria has been well studied, this branch is less understood in gram+ bacteria. in this study, corynebacterium glutamicum was cultivated with protocatechuate, p-cresol, vanillate and 4-hydroxybenzoate as sole carbon and energy sources for growth. enzymatic assays indicated that growing cells on these aromatic compounds exhibited protocatechuate 3,4-dioxygenase activities. data-mining of the genome of this bacterium re ...200516092756
the pep-pyruvate-oxaloacetate node as the switch point for carbon flux distribution in bacteria.in many organisms, metabolite interconversion at the phosphoenolpyruvate (pep)-pyruvate-oxaloacetate node involves a structurally entangled set of reactions that interconnects the major pathways of carbon metabolism and thus, is responsible for the distribution of the carbon flux among catabolism, anabolism and energy supply of the cell. while sugar catabolism proceeds mainly via oxidative or non-oxidative decarboxylation of pyruvate to acetyl-coa, anaplerosis and the initial steps of gluconeoge ...200416102602
e1 enzyme of the pyruvate dehydrogenase complex in corynebacterium glutamicum: molecular analysis of the gene and phylogenetic aspects.the e1p enzyme is an essential part of the pyruvate dehydrogenase complex (pdhc) and catalyzes the oxidative decarboxylation of pyruvate with concomitant acetylation of the e2p enzyme within the complex. we analyzed the corynebacterium glutamicum acee gene, encoding the e1p enzyme, and constructed and characterized an e1p-deficient mutant. sequence analysis of the c. glutamicum acee gene and adjacent regions revealed that acee is not flanked by genes encoding other enzymes of the pdhc. transcrip ...200516109942
identification and characterization of porh, a new cell wall channel of corynebacterium glutamicum.the cell wall of corynebacterium glutamicum contains the cation-selective channel (porin) pora(c.glut) and the anion-selective channel porb(c.glut) for the passage of hydrophilic solutes. lipid bilayer experiments with organic solvent extracts of whole c. glutamicum cells cultivated in minimal medium suggested that also another cation-selective channel-forming protein, named porh(c.glut), is present in c. glutamicum. the protein was purified to homogeneity by fast-protein liquid chromatography a ...200516112217
enhanced glutamic acid production by a h+-atpase-defective mutant of corynebacterium glutamicum.previously we reported that a mutant of corynebacterium glutamicum atcc14067 with reduced h+-atpase activity, f172-8, showed an approximately two times higher specific rate of glucose consumption than the parent, but no glutamic acid productivity under the standard biotin-limited culture conditions, where biotin concentration was set at 5.5 microg/l in the production medium (sekine et al., appl. microbiol. biotechnol., 57, 534-540 (2001)). in this study, various culture conditions were tested to ...200516116273
functional genomics and expression analysis of the corynebacterium glutamicum fpr2-cysixhdnyz gene cluster involved in assimilatory sulphate reduction.corynebacterium glutamicum is a high-gc gram-positive soil bacterium of great biotechnological importance for the production of amino acids. to facilitate the rational design of sulphur amino acid-producing strains, the pathway for assimilatory sulphate reduction providing the necessary reduced sulfur moieties has to be known. although this pathway has been well studied in gram-negative bacteria like escherichia coli and low-gc gram-positives like bacillus subtilis, little is known for the actin ...200516159395
the cgl2612 protein from corynebacterium glutamicum is a drug resistance-related transcriptional repressor: structural and functional analysis of a newly identified transcription factor from genomic dna analysis.the emergence of antibiotic-resistant bacteria often causes serious clinical problems. the tetr family is one of the major transcription factor families that regulate expression of genes involved in bacterial antimicrobial resistance systems. cgl2612 protein is a transcription factor newly identified by genomic dna analysis on corynebacterium glutamicum, which belongs to the mycolic acid-containing actinomycetales, including the well known pathogens corynebacterium diphtheriae and mycobacterium ...200516166084
the arac-type regulator ripa represses aconitase and other iron proteins from corynebacterium under iron limitation and is itself repressed by dtxr.the mrna level of the aconitase gene acn of corynebacterium glutamicum is reduced under iron limitation. here we show that an arac-type regulator, termed ripa for "regulator of iron proteins a," is involved in this type of regulation. a c. glutamicum deltaripa mutant has a 2-fold higher aconitase activity than the wild type under iron limitation, but not under iron excess. comparison of the mrna profiles of the deltaripa mutant and the wild type revealed that the acn mrna level was increased in ...200516179344
metabolic engineering of corynebacterium glutamicum for fuel ethanol production under oxygen-deprivation conditions.the central metabolic pathway of corynebacterium glutamicum was engineered to produce ethanol. a recombinant strain which expressed the zymomonas mobilis genes coding for pyruvate decarboxylase (pdc) and alcohol dehydrogenase (adhb) was constructed. both genes placed under the control of the c. glutamicum ldha promoter were expressed at high levels in c. glutamicum, resulting, under oxygen-deprivation conditions, in a significant yield ofethanol from glucose in a process characterized by the abs ...200416179801
the transcriptional regulator ssur activates expression of the corynebacterium glutamicum sulphonate utilization genes in the absence of sulphate.in a recent study, the putative regulatory gene cg0012 was shown to belong to the regulon of mcbr, a global transcriptional regulator of sulphur metabolism in corynebacterium glutamicum atcc 13032. a deletion of cg0012, now designated ssur (sulphonate sulphur utilization regulator), led to the mutant strain c. glutamicum dk100, which was shown to be blocked in the utilization of sulphonates as sulphur sources. according to dna microarray hybridizations, transcription of the ssu and seu genes, en ...200516194234
regulation of amtr-controlled gene expression in corynebacterium glutamicum: mechanism and characterization of the amtr regulon.amtr, the master regulator of nitrogen control in corynebacterium glutamicum, represses transcription of a number of genes during nitrogen surplus. repression is released by an interaction of amtr with signal transduction protein glnk. as shown by pull-down assays and gel retardation experiments, only adenylylated glnk, which is present in the cells during nitrogen limitation, is able to bind to amtr. the amtr regulon was characterized in this study by a combination of bioinformatics, transcript ...200516194241
biotechnological production of amino acids and derivatives: current status and prospects.for almost 50 years now, biotechnological production processes have been used for industrial production of amino acids. market development has been particularly dynamic for the flavor-enhancer glutamate and the animal feed amino acids l: -lysine, l: -threonine, and l: -tryptophan, which are produced by fermentation processes using high-performance strains of corynebacterium glutamicum and escherichia coli from sugar sources such as molasses, sucrose, or glucose. but the market for amino acids in ...200516195792
characterization of a corynebacterium glutamicum lactate utilization operon induced during temperature-triggered glutamate production.gene expression changes of glutamate-producing corynebacterium glutamicum were identified in transcriptome comparisons by dna microarray analysis. during glutamate production induced by a temperature shift, c. glutamicum strain 2262 showed significantly higher mrna levels of the ncgl2816 and ncgl2817 genes than its non-glutamate-producing derivative 2262np. reverse transcription-pcr analysis showed that the two genes together constitute an operon. ncgl2816 putatively codes for a lactate permease ...200516204505
role of the ssu and seu genes of corynebacterium glutamicum atcc 13032 in utilization of sulfonates and sulfonate esters as sulfur sources.corynebacterium glutamicum atcc 13032 was found to be able to utilize a broad range of sulfonates and sulfonate esters as sulfur sources. the two gene clusters potentially involved in sulfonate utilization, ssud1cba and ssui-seuabc-ssud2, were identified in the genome of c. glutamicum atcc 13032 by similarity searches. while the ssu genes encode proteins resembling ssu proteins from escherichia coli or bacillus subtilis, the seu gene products exhibited similarity to the dibenzothiophene-degradin ...200516204527
analysis of genes involved in arsenic resistance in corynebacterium glutamicum atcc 13032.corynebacterium glutamicum is able to grow in media containing up to 12 mm arsenite and 500 mm arsenate and is one of the most arsenic-resistant microorganisms described to date. two operons (ars1 and ars2) involved in arsenate and arsenite resistance have been identified in the complete genome sequence of corynebacterium glutamicum. the operons ars1 and ars2 are located some distance from each other in the bacterial chromosome, but they are both composed of genes encoding a regulatory protein ( ...200516204540
the whce gene of corynebacterium glutamicum is important for survival following heat and oxidative stress.in this study, we have analyzed an orf from corynebacterium glutamicum, which codes for a homologue of the streptomyces coelicolor whib-family of proteins known to be involved in sporulation. this orf encoded a putative protein which harbors a helix-turn-helix dna-binding motif and a probable redox-sensing motif, and has been designated whce. we constructed a whce mutant strain and analyzed the strain under a variety of growth conditions. this mutant strain exhibited a prolonged lag phase and ea ...200516212936
enhancement of biomolecule transport by electroporation: a review of theory and practical application to transformation of corynebacterium glutamicum.selective and reversible permeabilization of the cell wall permeability barrier is the focus for many biotechnological applications. in this article, the basic principles for reversible membrane permeabilization, based on biological, chemical, and physical methods are reviewed. emphasis is given to electroporation (electropermeabilization) which tends to be the most popular method for membrane permeabilization and for introduction of foreign molecules into the cells. the applications of this met ...200616224791
redirection of carbon flux to lysine in a recombinant of corynebacterium lactofermentum atcc 21799 by limited supply of pantothenate.to increase carbon flux to lysine, minimized production of amino acids that are biosynthetically related to lysine, for example, isoleucine and valine, is required. by limiting the supply of pantothenate, the precursor of coenzyme a, the carbon flux was redirected from isoleucine and valine to lysine in the recombinant of corynebacterium lactofermentum atcc 21799 containing the plasmid pgc77. the pgc77 contains hom(dr), thrb, and ilva encoding feedback-deregulated homoserine dehydrogenase, homos ...199916232592
development of a kinetic model for l-lysine biosynthesis in corynebacterium glutamicum and its application to metabolic control analysis.a mathematical model describing intracellular lysine synthesis by corynebacterium glutamicum in batch fermentation was developed. the model is based on material balance equations of the key metabolites, and includes mechanistically based, experimentally matched rate equations for individual enzymes. from the measurements of the levels of intra- and extracellular metabolites during cultivation, the kinetic parameters in the model were identified through the decomposition of the network of reactio ...199916232634
metabolic control analysis for lysine synthesis using corynebacterium glutamicum and experimental verification.metabolic control analysis was carried out for the lysine biosynthetic pathway in corynebacterium glutamicum atcc 21253 using mechanism-based kinetic models developed for enzymatic reactions for this pathway. the rate-limiting steps during lysine production were determined with the aid of flux control coefficients, which indicated that the lysine synthesis flux is governed mainly by both aspartokinase and permease activity in the export system. analyses indicated that an increase in the activity ...200016232840
attenuation control of ilvbnc in corynebacterium glutamicum: evidence of leader peptide formation without the presence of a ribosome binding site.the ilvbnc operon of corynebacterium glutamicum encodes acetohydroxy acid synthase and isomero-reductase, which are key enzymes of l-isoleucine, l-valine and l-leucine syntheses. in this study we identified the transcript initiation site of ilvbnc operon 292 nucleotides in front of the first structural gene, and detected the formation of a short transcript from the leader region in addition to the full-length transcript of the operon. this identifies the control of ilvbnc transcription by an att ...200016232899
effect of capsule circulation velocity on production of l-lysine by encapsulated corynebacterium glutamicum in an airlift bioreactor.the effect of capsule circulation velocity and volumetric oxygen transfer coefficient on the production of l-lysine by encapsulated corynebacterium glutamicum in an airlift bioreactor has been evaluated. a larger oxygen supply in the airlift bioreactor caused a more than 58% increase in l-lysine productivity compared to that in a shaking flask incubator. the quantity of l-lysine produced during 5 h of cultivation in the airlift bioreactor was suggested to increase with increasing circulation vel ...200116232951
triggering mechanism of l-glutamate overproduction by dtsr1 in coryneform bacteria.the mechanism of l-glutamate-overproduction by corynebacterium glutamicum, a biotin auxotroph, is very unique and interesting. l-glutamate overproduction by this bacterium is induced by biotin-limitation and suppressed by an excess of biotin. addition of a surfactant or penicillin is also induces l-glutamate overproduction even under excess biotin. after the development of general molecular biological tools such as cloning vectors and dna transfer techniques, genes encoding biosynthetic enzymes ...200216233348
purification and characterization of malate dehydrogenase from corynebacterium glutamicum.the malate dehydrogenase (mdh) (ec 1.1.1.37) from corynebacterium glutamicum (brevibacterium flavum) atcc14067 was purified to homogeneity. its amino-terminal sequence (residues 1 to 8) matched the sequence (residues 2 to 9) of the mdh from c. glutamicum (genbank accession no. cac83073). the molecular mass of the native enzyme was 130 kda. the protein was a homotetramer, with a 33-kda subunit molecular mass. the enzyme was almost equally active both for nadu and nadph as coenzyme on the bases of ...200316233457
characterization of bacterial drug antiporters homologous to mammalian neurotransmitter transporters.multidrug transporters are ubiquitous proteins, and, based on amino acid sequence similarities, they have been classified into several families. here we characterize a cluster of archaeal and bacterial proteins from the major facilitator superfamily (mfs). one member of this family, the vesicular monoamine transporter (vmat) was previously shown to remove both neurotransmitters and toxic compounds from the cytoplasm, thereby conferring resistance to their effects. a blast search of the available ...200516237035
the osmotic activation of transporter prop is tuned by both its c-terminal coiled-coil and osmotically induced changes in phospholipid composition.transporter prop of escherichia coli (propec) senses extracellular osmolality and mediates osmoprotectant uptake when it is rising or high. a replica of the propec c terminus (asp468-arg497) forms an intermolecular alpha-helical coiled-coil. this structure is implicated in the osmoregulation of intact propec, in vivo. like that from corynebacterium glutamicum (propcg), the prop orthologue from agrobacterium tumefaciens (propat) sensed and responded to extracellular osmolality after expression in ...200516239220
pyridinium-based ionic liquid matrices can improve the identification of proteins by peptide mass-fingerprint analysis with matrix-assisted laser desorption/ionization mass spectrometry.matrix-assisted laser desorption/ionization mass spectrometry has become an indispensable tool for identification of proteins by peptide mass-fingerprint analysis. selection of the matrix, addition of matrix additives, and sample-preparation techniques are known to affect the quality of the spectra and hence protein identification. we investigated the effect of pyridine as matrix additive for the commonly used crystalline matrix alpha-cyano-4-hydroxycinnamic acid (cca), forming a pyridinium base ...200616252087
functional analysis of all aminotransferase proteins inferred from the genome sequence of corynebacterium glutamicum.twenty putative aminotransferase (at) proteins of corynebacterium glutamicum, or rather pyridoxal-5'-phosphate (plp)-dependent enzymes, were isolated and assayed among others with l-glutamate, l-aspartate, and l-alanine as amino donors and a number of 2-oxo-acids as amino acceptors. one outstanding at identified is alat, which has a broad amino donor specificity utilizing (in the order of preference) l-glutamate > 2-aminobutyrate > l-aspartate with pyruvate as acceptor. another at is avta, which ...200516267288
metabolic engineering of corynebacterium glutamicum for l-serine production.although l-serine proceeds in just three steps from the glycolytic intermediate 3-phosphoglycerate, and as much as 8% of the carbon assimilated from glucose is directed via l-serine formation, previous attempts to obtain a strain producing l-serine from glucose have not been successful. we functionally identified the genes serc and serb from corynebacterium glutamicum, coding for phosphoserine aminotransferase and phosphoserine phosphatase, respectively. the overexpression of these genes, togeth ...200516269752
[metabolic flux analysis of l-tryptophan biosynthesis].the metabolic flux balance model of l-try synthesis by corynebacterium glutamicum was constructed in this paper. using this model, the metabolic flux distribution during the middle and late period were determined and the optimal flux distribution were calculated by linear program of matlab software. the analysis results indicate that 24.85% metabolic flux entered the hmp pathway and 75.15% entered the emp cycle. but comparing to the optimal flux distributions, the production of l-try should be i ...200316276922
toward the complete membrane proteome: high coverage of integral membrane proteins through transmembrane peptide detection.to attain a comprehensive membrane proteome of two strains of corynebacterium glutamicum (l-lysine-producing and the characterized model strains), both sample pretreatment and analysis methods were optimized. isolated bacterial membranes were digested with trypsin/cyanogen bromide or trypsin/chymotrypsin, and a complementary protein set was identified using the multidimensional protein identification technology (mudpit). besides a distinct number of cytosolic or membrane-associated proteins, the ...200616291997
the cytosolic, cell surface and extracellular proteomes of the biotechnologically important soil bacterium corynebacterium efficiens ys-314 in comparison to those of corynebacterium glutamicum atcc 13032.reference maps of the cytosolic, cell surface and extracellular proteome fractions of the amino acid-producing soil bacterium corynebacterium efficiens ys-314 were established. the analysis window covers a pi range from 3 to 7 along with a molecular mass range from 10 to 130 kda. after second-dimensional separation on sds-page and coomassie staining, computational analysis detected 635 protein spots in the cytosolic proteome fraction, whereas 76 and 102 spots were detected in the cell surface an ...200616302278
fluorescent phospholipid analogs as microscopic probes for detection of the mycolic acid-containing layer in corynebacterium glutamicum: detecting alterations in the mycolic acid-containing layer following ethambutol treatment.corynebacterium glutamicum belongs to the mycolic acid-containing actinomycetes, which also include mycobacterium, nocardia, and rhodococcus. the cells of this group possess a cell wall with a thick outer layer composed primarily of mycolic acid, which functions as a permeability barrier. to investigate the mechanism of mycolic acid-containing layer (mycolate layer) formation, we have developed a fluorescence microscopic technique detecting the mycolate layer in situ. the staining specificity of ...200516306684
the clgr protein regulates transcription of the clpp operon in bifidobacterium breve ucc 2003.five clp genes (clpc, clpb, clpp1, clpp2, and clpx), representing chaperone- and protease-encoding genes, were previously identified in bifidobacterium breve ucc 2003. in the present study, we characterize the b. breve ucc 2003 clpp locus, which consists of two paralogous genes, designated clpp1 and clpp2, whose deduced protein products display significant similarity to characterized clpp peptidases. transcriptional analyses showed that the clpp1 and clpp2 genes are transcribed in response to mo ...200516321946
class 1 integrons and tetracycline resistance genes in alcaligenes, arthrobacter, and pseudomonas spp. isolated from pigsties and manured soil.the presence of tetracycline resistance (tc(r)) genes and class i integrons (in-1), and their ability to cotransfer were investigated in tc(r) gram-negative (185 strains) and gram-positive (72 strains) bacteria from danish farmland and pigsties. the isolates belonged to the groups or species escherichia coli, enterobacter spp., arthrobacter spp., alcaligenes spp., pseudomonas spp., and corynebacterium glutamicum. the 257 isolates were screened for in-1. eighty-one of the gram-negative isolates w ...200516332771
new multiple-deletion method for the corynebacterium glutamicum genome, using a mutant lox sequence.due to the difficulty of multiple deletions using the cre/loxp system, a simple, markerless multiple-deletion method based on a cre/mutant lox system combining a right-element (re) mutant lox site with a left-element (le) mutant lox site was employed for large-scale genome rearrangements in corynebacterium glutamicum. eight distinct genomic regions that had been identified previously by comparative analysis of c. glutamicum r and c. glutamicum 13032 genomes were targeted for deletion. by homolog ...200516332837
amplified expression of fructose 1,6-bisphosphatase in corynebacterium glutamicum increases in vivo flux through the pentose phosphate pathway and lysine production on different carbon sources.the overexpression of fructose 1,6-bisphosphatase (fbpase) in corynebacterium glutamicum leads to significant improvement of lysine production on different sugars. amplified expression of fbpase via the promoter of the gene encoding elongation factor tu (eftu) increased the lysine yield in the feedback-deregulated lysine-producing strain c. glutamicum lyscfbr by 40% on glucose and 30% on fructose or sucrose. additionally formation of the by-products glycerol and dihydroxyacetone was significantl ...200516332851
microbial production of lysine and threonine from whey permeate.extracellular accumulation of lysine and threonine was investigated in modified whey permeate by using brevibacterium lactofermentum atcc 21086 and escherichia coli atcc 21151. whey permeate was prepared from whey by membrane ultrafiltration, and lactose was hydrolyzed by treating permeate with hcl or beta-galactosidase. the highest amount of lysine (3.3 g/liter) was produced from a mixture of acid-hydrolyzed whey permeate and yeast extract (0.2%). the highest amount of threonine (3.6 g/liter) w ...198316346209
improvement of an l-leucine-producing mutant of brevibacterium lactofermentum 2256 by genetically desensitizing it to alpha-acetohydroxy acid synthetase.genetic improvement of l-leucine productivity in strain 218, an ile 2-thiazolealanine-resistant mutant of brevibacterium lactofermentum 2256, was attempted. in strain 218, which produced 28 mg of l-leucine per ml from 13% glucose, alpha-isopropylmalate synthetase was genetically desensitized and derepressed to the effect of l-leucine, whereas alpha-acetohydroxy acid synthetase remained unaltered, although it could be derepressed phenotypically by limiting the isoleucine concentration in the cult ...198616347048
protoplast transformation in coryneform bacteria and introduction of an alpha-amylase gene from bacillus amyloliquefaciens into brevibacterium lactofermentum.[this corrects the article on p. 638 in vol. 51.].198616347093
microbiological degradation of malodorous substances of swine waste under aerobic conditions.phenol, p-cresol, and volatile fatty acids (vfa; acetic, propionic, isobutyric, butyric, isovaleric, and valeric acids) were used as odor indicators of swine waste. aeration of the waste allowed the indigenous microorganisms to grow and degrade these malodorous substances. the time required for degradation of these substances varied according to the waste used, and it was not necessarily related to their concentrations. using a minimal medium which contained one of the malodorous compounds as so ...198716347254
control of the lysine biosynthesis sequence in corynebacterium glutamicum as analyzed by overexpression of the individual corresponding genes.the gene cluster that codes for feedback-resistant aspartate kinase (lyscalpha and lyscbeta) and aspartate semialdehyde dehydrogenase (asd) was cloned from a mutant strain of corynebacterium glutamicum. its functional analysis by subcloning, enzyme assays, and type of aspartate kinase regulation enabled the isolation of a fragment for separate expression of the feedback-resistant kinase without aspartate semialdehyde dehydrogenase expression. this was used together with other clones constructed ...199116348510
metabolic engineering to produce tyrosine or phenylalanine in a tryptophan-producing corynebacterium glutamicum strain.the aromatic amino acids are synthesized via a common biosynthetic pathway. a tryptophan-producing mutant of corynebacterium glutamicum was genetically engineered to produce tyrosine or phenylalanine in abundance. to achieve this, three biosynthetic genes encoding the first enzyme in the common pathway, 3-deoxy-d-arabino-heptulosonate 7-phosphate synthase (ds), and the branch-point enzymes chorismate mutase and prephenate dehydratase were individually cloned from regulatory mutants of c. glutami ...199216348670
strains of corynebacterium glutamicum with different lysine productivities may have different lysine excretion systems.the lysine excretion systems of three different lysine-producing strains of corynebacterium glutamicum were characterized in intact cells. two strains (dg 52-5 and mh 20-22b) are lysine producers of different efficiency. they were bred by classical mutagenesis and have a feedback-resistant aspartate kinase. the third strain (kk 25) was constructed from the wild type by introducing the feedback-resistant aspartate kinase gene of strain mh 20-22b into its genome. the three strains were shown to po ...199316348855
on-line detection of substrate exhaustion by using nad(p)h fluorescence.in this study we examined the utility of nad(p)h fluorescence for monitoring aerobic fermentations of the threonine auxotroph corynebacterium glutamicum atcc 14296. instead of attempting complicated mathematical corrections for inner-filter effects, we found that it is possible to use the information contained in the on-line nad(p)h fluorescence signal to assess culture metabolic activities during fermentation. the first derivative of the filtered fluorescence signal, which approximates the turn ...199316348878
glutamate dehydrogenase is not essential for glutamate formation by corynebacterium glutamicum.two corynebacterium glutamicum strains, one being glutamate dehydrogenase (gdh) negative and the other possessing 11-fold-higher specific gdh activity than the parental wild type, were constructed and used to analyze the role of gdh in c. glutamicum. the results indicate (i) that gdh is dispensable for glutamate synthesis required for growth and (ii) that although a high level of gdh increases the intracellular glutamate pool, the level of gdh has no influence on glutamate secretion.199316349003
stable expression of hom-1-thrb in corynebacterium glutamicum and its effect on the carbon flux to threonine and related amino acids.the hom-1-thrb operon encodes homoserine dehydrogenase resistant to feedback inhibition by l-threonine and homoserine kinase. stable expression of this operon has not yet been attained in different corynebacterium glutamicum strains. we studied the use of chromosomal integration and of a low-copy-number vector for moderate expression of the hom-1-thrb operon to enable an analysis of the physiological consequences of its expression in c. glutamicum. strains carrying one, two, or three copies of h ...199416349146
efficient transformation of the cephamycin c producer nocardia lactamdurans and development of shuttle and promoter-probe cloning vectors.a high transformation efficiency (1 x 10 to 7 x 10 transformants per mug of dna) of nocardia lactamdurans lc411 was obtained by direct treatment of mycelium with polyethylene glycol 1000 and cesium chloride. a variety of vectors from streptomyces lividans, brevibacterium lactofermentum, rhodococcus fascians, and a nocardia (amycolatopsis) sp. were tested; transformants could be obtained only with vectors derived from an endogenous plasmid of the amycolatopsis sp. strain dsm 43387. vectors carryi ...199416349436
multiarray sensors with pattern recognition for the detection, classification, and differentiation of bacteria at subspecies and strain levels.this work describes the integration of a fully autonomous electrochemical biosensor with pattern recognition techniques for the detection and classification of bacteria at subspecies and strain level. the system provides a continuous, real-time monitoring of bacteria activity upon exposure to antibiotics. the system utilizes 96-well-type electrodes array (dox-dissolved oxygen sensor) with principal component analysis (pca) for rapid and routine classification of different classes of bacteria and ...200516351141
quantification of s-adenosyl methionine in microbial cell extracts.a sensitive method for quantification of s-adenosyl methionine (sam) in microbial cell extracts was developed and applied to corynebacterium glutamicum. the method is based on sam being completely hydrolyzed into (18)o-homoserine when extracted in boiling h(2) (18)o and thus can be clearly distinguished by gc-ms analysis from naturally labeled homoserine present in the cell extract. additional quantification of the total homoserine pool, representing both sam and homoserine, via hplc allows sepa ...200616369687
towards bacterial strains overproducing l-tryptophan and other aromatics by metabolic engineering.the aromatic amino acids, l-tryptophan, l-phenylalanine, and l-tyrosine, can be manufactured by bacterial fermentation. until recently, production efficiency of classical aromatic amino-acid-producing mutants had not yet reached a high level enough to make the fermentation method the most economic. with the introduction of recombinant dna technology, it has become possible to apply more rational approaches to strain improvement. many recent activities in this metabolic engineering have led to se ...200616374633
characterization and use of catabolite-repressed promoters from gluconate genes in corynebacterium glutamicum.the genes involved in gluconate catabolism (gntp and gntk) in corynebacterium glutamicum are scattered in the chromosome, and no regulatory genes are apparently associated with them, in contrast with the organization of the gnt operon in escherichia coli and bacillus subtilis. in c. glutamicum, gntp and gntk are essential genes when gluconate is the only carbon and energy source. both genes contain upstream regulatory regions consisting of a typical promoter and a hypothetical cyclic amp (camp) ...200616385030
accumulation of homolanthionine and activation of a novel pathway for isoleucine biosynthesis in corynebacterium glutamicum mcbr deletion strains.in the present work, the metabolic consequences of the deletion of the methionine and cysteine biosynthesis repressor protein (mcbr) in corynebacterium glutamicum, which releases almost all enzymes of methionine biosynthesis and sulfate assimilation from transcriptional regulation (d. a. rey, a. pühler, and j. kalinowski, j. biotechnol. 103:51-65, 2003), were studied. c. glutamicum atcc 13032 deltamcbr showed no overproduction of methionine. metabolome analysis revealed drastic accumulation of a ...200616385051
two-component systems of corynebacterium glutamicum: deletion analysis and involvement of the phos-phor system in the phosphate starvation response.corynebacterium glutamicum contains genes for 13 two-component signal transduction systems. in order to test for their essentiality and involvement in the adaptive response to phosphate (pi) starvation, a set of 12 deletion mutants was constructed. one of the mutants was specifically impaired in its ability to grow under pi limitation, and therefore the genes lacking in this strain were named phos (encoding the sensor kinase) and phor (encoding the response regulator). dna microarray analyses wi ...200616385062
transcriptional analysis of the f0f1 atpase operon of corynebacterium glutamicum atcc 13032 reveals strong induction by alkaline ph.corynebacterium glutamicum, a soil gram-positive bacterium used for industrial amino acid production, was found to grow optimally at ph 7.0-9.0 when incubated in 5 litre fermenters under ph-controlled conditions. the highest biomass was accumulated at ph 9.0. growth still occurred at ph 9.5 but at a reduced rate. the expression of the ph-regulated f0 f1 atpase operon (containing the eight genes atpbefhagdc) was induced at alkaline ph. a 7.5 kb transcript, corresponding to the eight-gene operon, ...200616385111
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