Large real-time holographic 3D displays: enabling components and results.

A holographic 3D display with 300  mm×200  mm active area was built. The display includes a spatial light modulator that modulates amplitude and phase of light and thus enables holographic reconstruction with high efficiency. Furthermore, holographic optical elements in photopolymer films and laser light sources are used.

Parkinson disease-associated mutation R1441H in LRRK2 prolongs the "active state" of its GTPase domain.

Mutation in leucine-rich-repeat kinase 2 (LRRK2) is a common cause of Parkinson disease (PD). A disease-causing point mutation R1441H/G/C in the GTPase domain of LRRK2 leads to overactivation of its kinase domain. However, the mechanism by which this mutation alters the normal function of its GTPase domain [Ras of complex proteins (Roc)] remains unclear.

Combined fluxomics and transcriptomics analysis of glucose catabolism via a partially cyclic pentose phosphate pathway in Gluconobacter oxydans 621H.

In this study, the distribution and regulation of periplasmic and cytoplasmic carbon fluxes in Gluconobacter oxydans 621H with glucose were studied by (13)C-based metabolic flux analysis ((13)C-MFA) in combination with transcriptomics and enzyme assays. For (13)C-MFA, cells were cultivated with specifically (13)C-labeled glucose, and intracellular metabolites were analyzed for their labeling pattern by liquid chromatography-mass spectrometry (LC-MS). In growth phase I, 90% of the glucose was oxidized periplasmically to gluconate and partially further oxidized to 2-ketogluconate.

Influence of oxygen limitation, absence of the cytochrome bc(1) complex and low pH on global gene expression in Gluconobacter oxydans 621H using DNA microarray technology.

The genome-wide transcriptional responses of the strictly aerobic α-proteobacterium Gluconobacter oxydans 621H to oxygen limitation, to the absence of the cytochrome bc(1) complex, and to low pH were studied using DNA microarray analyses. Oxygen limitation caused expression changes of 486 genes, representing 20% of the chromosomal genes. Genes with an increased mRNA level included those for terminal oxidases, the cytochrome bc(1) complex, transhydrogenase, two alcohol dehydrogenases, heme biosynthesis, PTS proteins, proteins involved in cyclic diGMP synthesis and degradation, two sigma factors, flagella and chemotaxis proteins, several stress proteins, and a putative exporter protein.

Proteomic Characterization of Cerebrospinal Fluid from Ataxia-Telangiectasia (A-T) Patients Using a LC/MS-Based Label-Free Protein Quantification Technology.

Cerebrospinal fluid (CSF) has been used for biomarker discovery of neurodegenerative diseases in humans since biological changes in the brain can be seen in this biofluid. Inactivation of A-T-mutated protein (ATM), a multifunctional protein kinase, is responsible for A-T, yet biochemical studies have not succeeded in conclusively identifying the molecular mechanism(s) underlying the neurodegeneration seen in A-T patients or the proteins that can be used as biomarkers for neurologic assessment of A-T or as potential therapeutic targets. In this study, we applied a high-throughput LC/MS-based label-free protein quantification technology to quantitatively characterize the proteins in CSF samples in order to identify differentially expressed proteins that can serve as potential biomarker candidates for A-T.

Knock-down of superoxide dismutase 1 sensitizes cisplatin-resistant human ovarian cancer cells.

Background: Overexpression of superoxide dismutase 1 (SOD1) has been shown to be one of the factors involved in causing cisplatin resistance in ovarian cancer. Reduction of SOD1 expression is expected to restore, at least partially, cisplatin sensitivity in ovarian cancer chemotherapy. Here, we explored the potential of RNAi as a therapy for reversal of cisplatin resistance.

Metabolic engineering of Gluconobacter oxydans for improved growth rate and growth yield on glucose by elimination of gluconate formation.

Gluconobacter oxydans N44-1, an obligatory aerobic acetic acid bacterium, oxidizes glucose primarily in the periplasm to the end products 2-ketogluconate and 2,5-diketogluconate, with intermediate formation of gluconate. Only a minor part of the glucose (less than 10%) is metabolized in the cytoplasm after conversion to gluconate or after phosphorylation to glucose-6-phosphate via the only functional catabolic routes, the pentose phosphate pathway and the Entner-Doudoroff pathway. This unusual method of glucose metabolism results in a low growth yield.

Targeted quantitative analysis of superoxide dismutase 1 in cisplatin-sensitive and cisplatin-resistant human ovarian cancer cells.

Protein quantification in a complex protein mixture presents a daunting task in biochemical analysis. Antibody-based immunoassays are traditional methods for protein quantification. However, there are issues associated with accuracy and specificity in these assays, especially when the changes are small (e.

Improving d-mannitol productivity of Escherichia coli: impact of NAD, CO2 and expression of a putative sugar permease from Leuconostoc pseudomesenteroides.

The highly productive whole-cell biotransformation of D-fructose to D-mannitol with recombinant, resting cells of Escherichia coli BL21(DE3) requires the combined expression of mdh, fdh and glf which encode mannitol and formate dehydrogenases and a sugar facilitator, respectively. However, long-term stability of the system was restricted, possibly due to loss of the cofactor NAD, high concentrations of formate, formation of CO(2) affecting the internal pH of the cells, accumulation of high intracellular concentrations of D-mannitol, and export of D-mannitol. Downstream of the mdh gene of Leuconostoc pseudomesenteroides, we identified an open reading frame encoding for a putative mannitol permease.

Myo-inositol facilitators IolT1 and IolT2 enhance D-mannitol formation from D-fructose in Corynebacterium glutamicum.

Reduction of D-fructose to D-mannitol by whole-cell biotransformation with recombinant resting cells of Corynebacterium glutamicum ATCC13032 requires the coexpression of mdh and fdh, which encode mannitol and formate dehydrogenases, respectively. However, d-mannitol formation is limited by the uptake of d-fructose in its unphosphorylated form, because additional expression of the sugar facilitator from Zymomonas mobilis resulted in a significantly increased productivity. Here we identified similarities of the myo-inositol transporters IolT1 and IolT2 of C.

Glucose oxidation and PQQ-dependent dehydrogenases in Gluconobacter oxydans.

Gluconobacter oxydans is famous for its rapid and incomplete oxidation of a wide range of sugars and sugar alcohols. The organism is known for its efficient oxidation of D-glucose to D-gluconate, which can be further oxidized to two different keto-D-gluconates, 2-keto-D-gluconate and 5-keto-D-gluconate, as well as 2,5-di-keto-D-gluconate. For this oxidation chain and for further oxidation reactions, G.

Response of the central metabolism of Corynebacterium glutamicum to different flux burdens.

To evaluate the importance of reactions within the central metabolism under different flux burdens the fluxes within the pentose phosphate pathway (PPP), as well as the other reactions of the central metabolism, were intensively analyzed and quantitated. For this purpose, Corynebacterium glutamicum was grown with [1-(13)C]glucose to metabolic and isotopic steady state and the fractional enrichments in precursor metabolites (e.g.

Activity of exporters of Escherichia coli in Corynebacterium glutamicum, and their use to increase L-threonine production.

L-Threonine is an important biotechnological product and Corynebacterium glutamicum is able to synthesize and accumulate this amino acid to high intracellular levels. We here use four exporters of Escherichia coli and show that three of them operate in C. glutamicum, with RhtA and RhtC being the most effective.

Insights into the structural basis of substrate recognition by histidinol-phosphate aminotransferase from Corynebacterium glutamicum.

Histidinol-phosphate aminotransferase (HisC) is a pyridoxal 5'-phosphate-dependent enzyme that catalyzes the reversible transamination reaction between histidinol phosphate (His-P) and 2-oxoglutarate (O-Glu). The crystal structures of apo histidinol-phosphate aminotransferase from Corynebacterium glutamicum, of the internal PLP aldimine adduct and of a pyridoxamine 5-phosphate-enzyme complex were determined at resolutions of 2.2, 2.

An easy cloning and expression vector system for Gluconobacter oxydans.

Gluconobacter oxydans is known for causing rapid and incomplete oxidation of a wide range of sugars, sugar acids and sugar alcohols. Therefore, this microorganism is already employed in several biotechnological processes that involve incomplete oxidation of a substrate, e.g.

D-mannitol production by resting state whole cell biotrans-formation of D-fructose by heterologous mannitol and formate dehydrogenase gene expression in Bacillus megaterium.

An in vivo system was developed for the biotransformation of D-fructose into D-mannitol by the expression of the gene mdh encoding mannitol dehydrogenase (MDH) from Leuconostoc pseudomesenteroides ATCC12291 in Bacillus megaterium. The NADH reduction equivalents necessary for MDH activity were regenerated via the oxidation of formate to carbon dioxide by coexpression of the gene fdh encoding Mycobacterium vaccae N10 formate dehydrogenase (FDH). High-level protein production of MDH in B.

The two carboxylases of Corynebacterium glutamicum essential for fatty acid and mycolic acid synthesis.

The suborder Corynebacterianeae comprises bacteria like Mycobacterium tuberculosis and Corynebacterium glutamicum, and these bacteria contain in addition to the linear fatty acids, unique alpha-branched beta-hydroxy fatty acids, called mycolic acids. Whereas acetyl-coenzyme A (CoA) carboxylase activity is required to provide malonyl-CoA for fatty acid synthesis, a new type of carboxylase is apparently additionally present in these bacteria. It activates the alpha-carbon of a linear fatty acid by carboxylation, thus enabling its decarboxylative condensation with a second fatty acid to afford mycolic acid synthesis.

Identification of a novel arabinofuranosyltransferase AftB involved in a terminal step of cell wall arabinan biosynthesis in Corynebacterianeae, such as Corynebacterium glutamicum and Mycobacterium tuberculosis.

Arabinofuranosyltransferase enzymes, such as EmbA, EmbB, and AftA, play pivotal roles in the biosynthesis of arabinogalactan, and the anti-tuberculosis agent ethambutol (EMB) targets arabinogalactan biosynthesis through inhibition of Mt-EmbA and Mt-EmbB. Herein, we describe the identification and characterization of a novel arabinofuranosyltransferase, now termed AftB (Rv3805c), which is essential in Mycobacterium tuberculosis. Deletion of its orthologue NCgl2780 in the closely related species Corynebacterium glutamicum resulted in a viable mutant.

Reduced folate supply as a key to enhanced L-serine production by Corynebacterium glutamicum.

The amino acid L-serine is required for pharmaceutical purposes, and the availability of a sugar-based microbial process for its production is desirable. However, a number of intracellular utilization routes prevent overproduction of L-serine, with the essential serine hydroxymethyltransferase (SHMT) (glyA) probably occupying a key position. We found that constructs of Corynebacterium glutamicum strains where chromosomal glyA expression is dependent on Ptac and lacIQ are unstable, acquiring mutations in lacIQ, for instance.

Topology and mutational analysis of the single Emb arabinofuranosyltransferase of Corynebacterium glutamicum as a model of Emb proteins of Mycobacterium tuberculosis.

The cell wall mycolyl-arabinogalactan (AG)--peptidoglycan complex is essential in mycobacterial species, such as Mycobacterium tuberculosis, and is the target of several antitubercular drugs. For instance, ethambutol (EMB) targets AG biosynthesis through inhibition of the arabinofuranosyltransferases Mt-EmbA and Mt-EmbB, as well as the single Emb from Corynebacterium glutamicum. Here, we present for the first time an experimental analysis of the membrane topology of Emb.

Characterization of myo-inositol utilization by Corynebacterium glutamicum: the stimulon, identification of transporters, and influence on L-lysine formation.

Although numerous bacteria possess genes annotated iol in their genomes, there have been very few studies on the possibly associated myo-inositol metabolism and its significance for the cell. We found that Corynebacterium glutamicum utilizes myo-inositol as a carbon and energy source, enabling proliferation with a high maximum rate of 0.35 h-1.

Modification of the membrane-bound glucose oxidation system in Gluconobacter oxydans significantly increases gluconate and 5-keto-D-gluconic acid accumulation.

Gluconobacter oxydans DSM 2343 (ATCC 621H)catalyzes the oxidation of glucose to gluconic acid and subsequently to 5-keto-D-gluconic acid (5-KGA), a precursor of the industrially important L-(+)-tartaric acid. To further increase 5-KGA production in G. oxydans, the mutant strain MF1 was used.

Arabinan-deficient mutants of Corynebacterium glutamicum and the consequent flux in decaprenylmonophosphoryl-D-arabinose metabolism.

The arabinogalactan (AG) of Corynebacterianeae is a critical macromolecule that tethers mycolic acids to peptidoglycan, thus forming a highly impermeable cell wall matrix termed the mycolyl-arabinogalactan peptidoglycan complex (mAGP). The front line anti-tuberculosis drug, ethambutol (Emb), targets the Mycobacterium tuberculosis and Corynebacterium glutamicum arabinofuranosyltransferase Mt-EmbA, Mt-EmbB and Cg-Emb enzymes, respectively, which are responsible for the biosynthesis of the arabinan domain of AG. The substrate utilized by these important glycosyltransferases, decaprenylmonophosphoryl-D-arabinose (DPA), is synthesized via a decaprenylphosphoryl-5-phosphoribose (DPPR) synthase (UbiA), which catalyzes the transfer of 5-phospho-ribofuranose-pyrophosphate (pRpp) to decaprenol phosphate to form DPPR.

High-yield 5-keto-D-gluconic acid formation is mediated by soluble and membrane-bound gluconate-5-dehydrogenases of Gluconobacter oxydans.

Gluconobacter oxydans DSM 2343 is known to catalyze the oxidation of glucose to gluconic acid, and subsequently, to 2-keto-D-gluconic acid (2-KGA) and 5-keto-D-gluconic acid (5-KGA), by membrane-bound and soluble dehydrogenases. In G. oxydans MF1, in which the membrane-bound gluconate-2-dehydrogenase complex was inactivated, formation of the undesired 2-KGA was absent.