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One of the most important industrial requirements is to find the materials which have the tendency to withstand harsh environments, such as high pressure, high temperature, and radiation environments, so that they will maintain high efficiency in these conditions. Such materials are greatly required for aerospace, defense, and high-temperature molecular device fabrications. In the present context, we have chosen l-isoleucine samples to assess the crystallographic phase stability under dynamic shocked conditions. The stability result has been screened by a powder x-ray diffractometer (PXRD). Based on the observed diffraction results, it is well evidenced by the cleavage planes, such as (002) (003), (004), (005), and (006), that the applied shock waves do not induce any crystallographic phase transitions and lattice deformations even at 200 shocks. The unit cell dimensions of the l-isoleucine sample were a = 9.752 Å, b = 5.321 Å, c = 14.13 Å and V = 728 Å3 which were found to be the same under all numbers of shocked conditions. Hence, the l-isoleucine sample is strongly suggested for device fabrication, and previous static high-pressure experimental results also support this suggestion. [ABSTRACT FROM AUTHOR]

As one of the three branched-chain amino acids essential for human body, L-isoleucine is widely used in food, medicine, and feed industries. At present, L-isoleucine is mainly produced by microbial fermentation, and the main production strain is Corynebacterium glutamicum. The biosynthetic pathway of L-isoleucine in C. glutamicum is complex, and the activity of key enzymes and the transcription of key genes in the pathway are strictly regulated. The intracellularly synthesized L-isoleucine is secreted by transporters, and the activity of the transporters is also regulated. These intricate regulatory mechanisms increase the difficulty to engineer the L-isoleucine-producing C. glutamicum. This article focuses on the mechanism of L-isoleucine biosynthesis, secretion, and regulation in C. glutamicum and reviews the various metabolic engineering strategies for improving L-isoleucine production efficiency in C. glutamicum. [ABSTRACT FROM AUTHOR]

Gout and hyperuricemia are highly prevalent metabolic diseases caused by high level of uric acid. Amino acids (AAs) involve in various biochemical processes including the biosynthesis of uric acid. However, the role of AAs in discriminating gout from hyperuricemia remains unknown. Here, we report that the plasma AAs profile can distinguish acute gout (AG) from asymptomatic hyperuricemia (AHU). We established an LC-MS/MS-based method to measure the plasma AAs without derivatization for the AG and AHU patients, and healthy controls. We found that the plasma profiling of AAs separated the AG patients from AHU patients and controls visually in both principal component analysis and orthogonal partial least-squares discriminant analysis (OPLS-DA) models. In addition, L-isoleucine, L-lysine, and L-alanine were suggested as the key mediators to distinguish the AG patients from AHU and control groups based on the S-plot analysis and variable importance in the projection values in the OPLS-DA models, volcano plot, and the receiver operating characteristic curves. In addition, the saturation of monosodium urate in the AA solutions at physiologically mimic status supported the changes in plasma AAs facilitating the precipitation of monosodium urate. This study suggests that L-isoleucine, L-lysine, and L-alanine could be the potential markers to distinguish the AG from AHU when the patients have similar blood levels of uric acid, providing new strategies for the prevention, treatment, and management of acute gout. [ABSTRACT FROM AUTHOR]

The experimental saturated solubilities of L-isoleucine, L-threonine, and L-serine in aqueous mixtures of a KCl solution at 298.15 K are presented in this article. The solubilities are measured by gravimetric method. In the present study the theoretical calculation of the standard transfer Gibbs free energy, cavity forming enthalpy of transfer, cavity forming transfer Gibbs free energy, dipole-dipole interaction effect have been computed. The chemical effects of the transfer Gibbs energies for the present amino acids have been obtained by subtracting the cavity effects and dipole-dipole interaction effects from the Δ G ( i). The stability of the experimental amino acids in aqueous KCl in terms of thermodynamic parameters is explained. [ABSTRACT FROM AUTHOR]

Corynebacterium glutamicum ssp. lactofermentum strain JHI3-156 produces l-isoleucine (Ile). Overexpression of the Saccharomyces cerevisiae-derived NADH kinase gene ( POS5) and the endogenous glucose-6-phosphate dehydrogenase and NAD kinase genes ( zwf- ppnK) in JHI3-156 increased Ile production by 26 and 31 %, respectively. To decipher the global effect of POS5 and zwf- ppnK overexpression on Ile biosynthesis, proteomic analysis was conducted. Twenty-four differentially expressed proteins were identified in the POS5-overexpressing strain, most of which are related to inositol catabolism, central carbon metabolism, anaplerotic pathway, protein biosynthesis and the stress response. In the zwf- ppnK-overexpressing strain, seven differentially-expressed proteins, including PpnK and anaplerotic enzymes, were identified. This result indicates the involvement of a novel inositol catabolism step and the importance of the anaplerotic pathway in Ile biosynthesis. This finding will be helpful in the systematic metabolic engineering of C. glutamicum for Ile biosynthesis. [ABSTRACT FROM AUTHOR]

Our previous work has shown that L-isoleucine production in Corynebacterium glutamicum IWJ001 could be increased by overexpressing ilvA1 encoding a feedback-resistant threonine dehydratase, ilvBN1 encoding a feedback-resistant acetohydroxy acid synthase, lrp encoding the global regulator Lrp, brnFE encoding the two-component export system BrnFE, or ppnk1 encoding NAD kinase. The main purpose of this study is to further increase the L-isoleucine production in C. glutamicum IWJ001 by overexpressing the above genes in various combinations. Several C. glutamicum strains IWJ001/pDXW-8- ppnk1- lrp-brnFE, IWJ001/pDXW-8- ilvBN1- ilvA1- lrp-brnFE, IWJ001/pDXW-8- ilvBN1- ilvA1- ppnk1, and IWJ001/pDXW-8- ppnk1- ilvBN1- ilvA1- lrp-brnFE were constructed, and L-isoleucine production and activities of several key enzymes in these strains were analyzed. Compared with the control strain IWJ001/pDXW-8, L-isoleucine production increased in all of the four strains. IWJ001/pDXW-8- ilvBN1- ilvA1- ppnk1 showed the highest L-isoleucine production and produced 32.3 g/L L-isoleucine in 72 h fed batch fermentation. The results indicate that L-isoleucine production in C. glutamicum could be increased by enhancing the carbon flux and NADPH supply in the biosynthetic pathway. [ABSTRACT FROM AUTHOR]

Corynebacterium glutamicum is the workhorse for the production of amino acids, including l-isoleucine (Ile). During Ile biosynthesis, NADPH is required as a crucial cofactor. In this study, four NADPH-supplying strategies based on NAD kinase, NADH kinase, glucose-6-phosphate dehydrogenase, and NAD kinase coupling with glucose-6-phosphate dehydrogenase were compared, and their influences on Ile biosynthesis were examined. PpnK is a NAD kinase of C. glutamicum ssp. lactofermentum JHI3-156 that predominantly phosphorylates NAD to produce NADP. Pos5 is a NADH kinase of Saccharomyces cerevisiae that predominantly phosphorylates NADH to produce NADPH. Zwf is a glucose-6-phosphate dehydrogenase of JHI3-156. The ppnK, POS5, zwf, and zwf- ppnK genes were overexpressed in the Ile-producing strain JHI3-156. The expression of all four genes increased intracellular NADPH concentration and Ile production. The increase of NADPH concentration and Ile production in a POS5-expressing strain (229 and 75.6 %, respectively) was higher than that in a ppnK-expression strain. The expression of zwf also increased NADPH supply and Ile biosynthesis, but the constitutive expression of zwf was not as effective as the inducible expression of zwf. Coexpression of zwf and ppnK genes greatly enhanced NADPH supply and thus improved Ile production by up to 85.9 %, indicating that this strategy was the most effective one. These results are helpful for improving Ile biosynthesis and other biosynthetic processes. [ABSTRACT FROM AUTHOR]

l-isoleucine is synthesized from 2-ketobutyrate and pyruvate in Corynebacterium glutamicum, and the supplies of these two precursors are important for l-isoleucine synthesis. C. glutamicum YILWΔ alaT with alaT gene deletion (encoding alanine aminotransferase, a principal enzyme for l-alanine synthesis) was constructed to increase intracellular pyruvate availability, and the thrABC genes from Escherichia coli (encoding bifunctional aspartate kinase I-homoserine dehydrogenase I, homoserine kinase, and threonine synthetase) were overexpressed in C. glutamicum YILW and YILWΔ alaT to increase the supply of intracellular 2-ketobutyrate. In the fed-batch fermentation, YILWpXMJ19 thrABC, YILWΔ alaT, and YILWΔ alaTpXMJ19 thrABC exhibited 5.3, 17.6, and 8.4 % higher l-isoleucine production than the original strain, respectively. Both YILWpXMJ19 thrABC and YILWΔ alaT excreted lower concentrations of l-lysine, l-alanine, and l-valine. YILWΔ alaTpXMJ19 thrABC exhibited a cumulative reduction of these by-products excretion, which indicated that thrABC overexpression combined with alaT deletion resulted in the metabolic flux redistribution from 2-ketobutyrate and pyruvate to l-isoleucine synthesis, and decreased the fluxes to by-products synthesis accordingly. [ABSTRACT FROM AUTHOR]

Biosynthetic threonine deaminase (TD) is a key enzyme for the synthesis of isoleucine which is allosterically inhibited and activated by Ile and Val, respectively. The binding sites of Ile and Val and the mechanism of their regulations in TD are not clear, but essential for a rational design of efficient productive strain(s) for Ile and related amino acids. In this study, structure-based computational approach and site-directed mutagenesis were combined to identify the potential binding sites of Ile and Val in Escherichia coli TD. Our results demonstrated that each regulatory domain of the TD monomer possesses two nonequivalent effector-binding sites. The residues R362, E442, G445, A446, Y369, I460, and S461 only interact with Ile while E347, G350, and F352 are involved not only in the Ile binding but also in the Val binding. By further considering enzyme kinetic data, we propose a concentration-dependent mechanism of the allosteric regulation of TD by Ile and Val. For the construction of Ile overproducing strain, a novel TD mutant with double mutation of F352A/R362F was also created, which showed both higher activity and much stronger resistance to Ile inhibition comparing to those of wild-type enzyme. Overexpression of this mutant TD in E. coli JW3591 significantly increased the production of ketobutyrate and Ile in comparison to the reference strains overexpressing wild-type TD or the catabolic threonine deaminase (TdcB). This work builds a solid basis for the reengineering of TD and related microorganisms for Ile production. [ABSTRACT FROM AUTHOR]

The β-defensins, expressed in epithelial cells of multiple tissues including intestine, play a critical role in the mammalian innate immunity. However, it is little known about the role of functional nutrients in the regulation of porcine β-defensins' expressions in intestinal epithelial cells. The present study was conducted to determine the hypothesis that zinc and l-isoleucine regulate the expressions of porcine β-defensins in IPEC-J2 cells. Cells were cultured in DMEM/F12 medium containing supplemental 0-500 μg/mL l-isoleucine or 0-500 μmol/mL zinc sulfate that was used to increase the concentration of Zn in the medium. At 12 h after the treatment by the appropriate concentrations of l-isoleucine or Zn, the mRNA and protein expressions of porcine β-defensin 1, 2 and 3 were increased ( P < 0.05), and reached their maximum after treatment with 25 or 100 μmol/mL zinc sulfate and 25 or 50 μg/mL isoleucine ( P < 0.05). These results suggested that both Zn and l-isoleucine could induce β-defensins' expressions in porcine intestinal epithelial cells. [ABSTRACT FROM AUTHOR]

In the present investigation, novel poly(amide-imide)/TiO bionanocomposites containing L-isoleucine moiety in the main chain were prepared via a simple and inexpensive ultrasonic irradiation process. Poly(amide-imide) as a source of polymer was synthesized by direct polycondensation reaction of N-trimellitylimido-L-isoleucine with 4,4′-diaminodiphenylsulfone in the tetrabutyl ammonium bromide/triphenyl phosphite as a green solvent system. Due to the high surface energy and tendency for agglomeration, the surface of TiO nanoparticles was modified with γ-aminopropyltriethoxyl silane. The resulting hybrid bionanocomposites were fully characterized by FT-IR spectroscopy, UV-vis spectroscopy, X-ray diffraction, field emission scanning electron microscopy, transmission electron microscopy (TEM) and thermogravimetric analysis. Through TEM photographs, it can be found that the surface modified TiO nanoparticles with diametric size of less than 40 nm, uniformly dispersed in the obtained PAI matrix. [ABSTRACT FROM AUTHOR]

New optically active poly(ester-imide)s PEIs were prepared from newly synthesized N,N′-(bicyclo[2,2,2]oct-7-ene-2,3,5,6-tetracarboxylic)-bis- l-isoleucine diacid 4 via direct polycondensation with various aromatic diols in a system of tosyl chloride (TsCl), pyridine (Py), and N,N-dimethylformamide (DMF). The reactions with bicyclo TsCl were significantly promoted by controlling alcoholysis with diols, in the presence of catalytic amounts of DMF, to give a series of optically active PEIs, with good yield and moderate inherent viscosity ranging from 0.44 to 0.66 dL/g. The diacid 4 was synthesized by the condensation reaction of [2,2,2]oct-7-ene-2,3,5,6-tetracarboxylic dianhydride 1 with l-isoleucine 2 in acetic acid. All of these polymers were highly organosoluble in solvents such as N,N-dimethylacetamide (DMAc), N,N-dimethylformamide (DMF), dimethylsulfoxide (DMSO), and N-methyl-2-pyrrolidone (NMP) at room temperature. They were fully characterized by means of NMR spectroscopy, FTIR spectroscopy, gel permeation chromatography (GPC), elemental analyses, inherent viscosity, solubility test, and specific rotation, and thermal properties of the poly(ester-imide)s were investigated using TGA/DTG. [ABSTRACT FROM AUTHOR]

The branched chain amino acids (BCAAs), l-valine, l-leucine, and l-isoleucine, have recently been attracting much attention as their potential to be applied in various fields, including animal feed additive, cosmetics, and pharmaceuticals, increased. Strategies for developing microbial strains efficiently producing BCAAs are now in transition toward systems metabolic engineering from random mutagenesis. The metabolism and regulatory circuits of BCAA biosynthesis need to be thoroughly understood for designing system-wide metabolic engineering strategies. Here we review the current knowledge on BCAAs including their biosynthetic pathways, regulations, and export and transport systems. Recent advances in the development of BCAA production strains are also reviewed with a particular focus on l-valine production strain. At the end, the general strategies for developing BCAA overproducers by systems metabolic engineering are suggested. [ABSTRACT FROM AUTHOR]

The female-produced sex pheromone of the scarab beetle Phyl- lophaga lanceolata was identified as the methyl ester of an essential amino acid, L-Ieucine. During field testing, 239 male P lanceolata were caught in traps baited with L-leucine methyl ester. L-Isoleucine and L-valine methyl esters, similar in structure to L-Ieucine methyl ester and previously identified as female-produced sex pheromone compounds employed by other Phyllophaga species, were also tested. Addition of L-valine or L-isoleucine methyl esters to the L-leucine methyl ester in 1:1 ratios completely inhibited attraction of P lanceolata males. Males of P squamipilosa were also captured using L-leucine methyl ester. This is the first record of P squamipilosa from Kansas. [ABSTRACT FROM AUTHOR]

The continuous administration of physiological doses of the branched-chain amino acids leucine, isoleucine, and valine (Leu-Ile-Val) to Yoshida sarcoma-bearing rats caused a significant increase in the survival time by 32% and a significant reduction of tumor size after 3 weeks of growth by 33%. The shift of the nitrogen balance to negative values during the cachectic stage was delayed but not prevented. On the average, less nitrogen (-47 mg/day) were lost by Leu-Ile-Val treated rats compared with untreated tumor-bearing animals (-91 mg N/day). It appeared that Leu-Ile-Val increased the synthesis of carcass proteins, while it left the proteolysis rate unchanged, since the excretion of urea and creatinine was unaffected by these amino acids. The daily excretion of α-ketoglutarate, which is correlated with tumor size during the early stage of growth, was decreased during the first 2 weeks by Leu-Ile-Val, but remained for a longer period on a high level than in untreated tumor bearers. The results point to an improvement of the metabolic resistance against carcass protein depletion of the tumor-bearing host by the administration of branched-chain amino acids. [ABSTRACT FROM AUTHOR]

Reactions of hydrochlorides of glycine, alanine, phenylalanine, L-isolcucinc, and L-valine esters with aromatic and heteroaromatic carboxamides afforded hydrochlorides of the corresponding N-(amidomethyl)-α-amino acid esters. N-(Phthalimidomethyl)-α-amino acid esters were obtained by reactions of α-amino acid esters containing free amino groups with formaldehyde and phthalimide, TheH NMR studies demonstrated that the chiral centers of α-amino acids may be retained in the course of condensation. Reactions of the Mannish bases obtained and their hydrochlorides with acetic anhydride and tosyl chloride afforded the corresponding N-aryl and N-sulfonyl derivatives. [ABSTRACT FROM AUTHOR]

A multienzyme system consisting of leucine dehydrogenase (EC 1.4.1.9), L-lactic dehydrogenase (EC 1.1.1.27), urease (EC 3.5.1.5), and dextran-NAD was microencapsulated within artificial cells. This system could convert ammonia and urea into essential amino acids, L-leucine, L-valine, and L-isoleucine. L-lactate acted as a cosubstrate for the regeneration of dextran-NADH. Greater concentrations of L-lactate favored the higher conversion ratios. The effects of ammonium salts and urea on reaction rate were also studied. The relative reaction rates in ammonium salts solutions were 44.6-78.8% of those in urea solutions. More than 90% of the original activity was retained when artificial cells were kept at 4°C for 6 wk. [ABSTRACT FROM AUTHOR]

L-glutamine dramatically augments insulin release evoked by L-leucine. The dynamics, specificity and environmental modifications of the insulin secretory response to the combination of L-glutamine and L-leucine were investigated. L-glutamine alone failed to stimulate proinsulin biosynthesis or insulin release in rat pancreatic islets, but augmented synthesis and secretion evoked by L-leucine. In relation to the secretory response, L-glutamine could be replaced by L-asparagine but not L-glutamate; L-leucine could be replaced by L-norvaline or L-isoleucine but not by L-valine, L-norleucine, glycine, L-serine, L-lysine, L-phenylalanine or L-arginine. The secretory response to L-leucine was rapid and biphasic, whereas the enhancing action of L-glutamine upon insulin release was progressive. The release of insulin evoked by the combination of L-leucine and L-glutamine was inhibited by antimycin A, menadione, ammonium ions, verapamil, adrenaline, and by the absence of extracellular Ca. It was increased at high carbon dioxide tension, and by glucose, theophylline and cytochalasin B. It is concluded that the enhancing action of L-glutamine upon insulin release evoked by certain amino acids represents a phenomenon dependent on the integrity of oxidative metabolism and involving essentially the same sequence of metabolic, ionic and motile events as that characterizing the process of glucose-induced insulin release. [ABSTRACT FROM AUTHOR]