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#28082680   2017/01/13 Save this To Up

Structural Determinants of the Gain-of-Function Phenotype of Human Leukemia-associated Mutant CBL Oncogene.

Mutations of the tyrosine kinase-directed ubiquitin ligase CBL cause myeloid leukemias, but the molecular determinants of the dominant leukemogenic activity of mutant CBL oncogenes are unclear. Here, we first define a gain-of-function attribute of the most common leukemia-associated CBL mutant, Y371H, by demonstrating its ability to increase proliferation of hematopoietic stem/progenitor cells (HSPCs) derived from CBL-null and CBL/CBL-B-null mice. Next, we express second-site point/deletion mutants of CBL-Y371H in CBL/CBL-B-null HSPCs or the cytokine-dependent human leukemic cell line TF-1 to show that individual or combined Tyr → Phe mutations of established phosphotyrosine residues (Tyr-700, Tyr-731, and Tyr-774) had little impact on the activity of the CBL-Y371H mutant in HSPCs, and the triple Tyr → Phe mutant was only modestly impaired in TF-1 cells. In contrast, intact tyrosine kinase-binding (TKB) domain and proline-rich region (PRR) were critical in both cell models. PRR deletion reduced the stem cell factor (SCF)-induced hyper-phosphorylation of the CBL-Y371H mutant and the c-KIT receptor and eliminated the sustained p-ERK1/2 and p-AKT induction by SCF. GST fusion protein pulldowns followed by phospho-specific antibody array analysis identified distinct CBL TKB domains or PRR-binding proteins that are phosphorylated in CBL-Y371H-expressing TF-1 cells. Our results support a model of mutant CBL gain-of-function in which mutant CBL proteins effectively compete with the remaining wild type CBL-B and juxtapose TKB domain-associated PTKs with PRR-associated signaling proteins to hyper-activate signaling downstream of hematopoietic growth factor receptors. Elucidation of mutant CBL domains required for leukemogenesis should facilitate targeted therapy approaches for patients with mutant CBL-driven leukemias.

1686 related Products with: Structural Determinants of the Gain-of-Function Phenotype of Human Leukemia-associated Mutant CBL Oncogene.

TCP-1 theta antibody Sour Recombinant Human GSTO1 M Recombinant Human GSTO1 M Recombinant Human GSTO1 M Recombinant Human T-cell Recombinant Human T-cell Recombinant Human T-cell Recombinant Human p53 Mut Recombinant Human p53 Mut Recombinant Human p53 Mut Recombinant Human PKC the Recombinant Human PKC the

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#27601900   2016/09/07 Save this To Up

The preosteoblast response of electrospinning PLGA/PCL nanofibers: effects of biomimetic architecture and collagen I.

Constructing biomimetic structure and incorporating bioactive molecules is an effective strategy to achieve a more favorable cell response. To explore the effect of electrospinning (ES) nanofibrous architecture and collagen I (COL I)-incorporated modification on tuning osteoblast response, a resorbable membrane composed of poly(lactic-co-glycolic acid)/poly(caprolactone) (PLGA/PCL; 7:3 w/w) was developed via ES. COL I was blended into PLGA/PCL solution to prepare composite ES membrane. Notably, relatively better cell response was delivered by the bioactive ES-based membrane which was fabricated by modification of 3,4-dihydroxyphenylalanine and COL I. After investigation by field emission scanning electron microscopy, Fourier transform infrared spectroscopy, contact angle measurement, and mechanical test, polyporous three-dimensional nanofibrous structure with low tensile force and the successful integration of COL I was obtained by the ES method. Compared with traditional PLGA/PCL membrane, the surface hydrophilicity of collagen-incorporated membranes was largely enhanced. The behavior of mouse preosteoblast MC3T3-E1 cell infiltration and proliferation on membranes was studied at 24 and 48 hours. The negative control was fabricated by solvent casting. Evaluation of cell adhesion and morphology demonstrated that all the ES membranes were more favorable for promoting the cell adhesion and spreading than the casting membrane. Cell Counting Kit-8 assays revealed that biomimetic architecture, surface topography, and bioactive properties of membranes were favorable for cell growth. Analysis of β1 integrin expression level by immunofluorescence indicated that such biomimetic architecture, especially COL I-grafted surface, plays a key role in cell adhesion and proliferation. The real-time polymerase chain reaction suggested that both surface topography and bioactive properties could facilitate the cell adhesion. The combined effect of biomimetic architecture with enhanced surface activity by 3,4-dihydroxyphenylalanine-assisted modification and COL I incorporation of PLGA/PCL electrospun membranes could successfully fill osteogenic defects and allow for better cell proliferation and differentiation.

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CAR,CAR,Constitutive acti Ofloxacin CAS Number [824 Picro-Sirius Red Stain K BACTERIOLOGY BACTEROIDES ELCI ELISA grade chick ty ELCI ELISA grade chick ty ELISA grade chick type I ELBI ELISA grade bovine t ELBI ELISA grade bovine t ELISA grade bovine type I ELISA grade porcine type ELPI ELISA grade porcine

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#26796258   2016/01/22 Save this To Up

The stimulatory effects of eNOS/F92A-Cav1 on NO production and angiogenesis in BMSCs.

Nitric oxide (NO) is generated in endothelial cells by endothelial nitric oxide synthase (eNOS). Caveolin-1 (Cav1) inhibits eNOS function and NO production. Modifying Cav1 scaffold domain, in particular Phenylalanine at position 92 (F92) is critical for the inhibitory actions of Cav1 toward eNOS. The aims of this study were to investigate the effect of enhanced NO production in term of in vitro angiogenesis on rat bone marrow derived mesenchymal stem cells (BMSCs) transduced with a novel bicistronic lentiviral vector co-expressing eNOS and mutant Cav1 (F92A).

1953 related Products with: The stimulatory effects of eNOS/F92A-Cav1 on NO production and angiogenesis in BMSCs.

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#26174624   2015/11/28 Save this To Up

Rapid metabolic analysis of Rhodococcus opacus PD630 via parallel 13C-metabolite fingerprinting.

For rapid analysis of microbial metabolisms, (13)C-fingerprinting employs a set of tracers to generate unique labeling patterns in key amino acids that can highlight active pathways. In contrast to rigorous (13)C-metabolic flux analysis ((13)C-MFA), this method aims to provide metabolic insights without expensive flux measurements. Using (13)C-fingerprinting, we investigated the metabolic pathways in Rhodococcus opacus PD630, a promising biocatalyst for the conversion of lignocellulosic feedstocks into value-added chemicals. Specifically, seven metabolic insights were gathered as follows: (1) glucose metabolism mainly via the Entner-Doudoroff (ED) pathway; (2) lack of glucose catabolite repression during phenol co-utilization; (3) simultaneous operation of gluconeogenesis and the ED pathway for the co-metabolism of glucose and phenol; (4) an active glyoxylate shunt in acetate-fed culture; (5) high flux through anaplerotic pathways (e.g., malic enzyme and phosphoenolpyruvate carboxylase); (6) presence of alternative glycine synthesis pathway via glycine dehydrogenase; and (7) utilization of preferred exogenous amino acids (e.g., phenylalanine). Additionally, a (13)C-fingerprinting kit was developed for studying the central metabolism of non-model microbial species. This low-cost kit can be used to characterize microbial metabolisms and facilitate the design-build-test-learn cycle during the development of microbial cell factories.

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Rapid Microplate Assay K Peroxide Block for Image Peroxide Block for Image Peroxide Block for Image H. Pylori Rapid Stain Ki H. Pylori Rapid Stain Ki Biotin Blocking Kit for Biotin Blocking Kit for Blue Feulgen DNA Ploidy H. Pylori Rapid Stain (1 H. Pylori Rapid Stain (1 Primary Antibody Dropper

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#26020360   2015/06/25 Save this To Up

Modeling the Histidine-Phenylalanine Interaction: The NH···π Hydrogen Bond of Imidazole·Benzene.

NH···π hydrogen bonds occur frequently between the amino acid side groups in proteins and peptides. Data-mining studies of protein crystals find that ∼80% of the T-shaped histidine···aromatic contacts are CH···π, and only ∼20% are NH···π interactions. We investigated the infrared (IR) and ultraviolet (UV) spectra of the supersonic-jet-cooled imidazole·benzene (Im·Bz) complex as a model for the NH···π interaction between histidine and phenylalanine. Ground- and excited-state dispersion-corrected density functional calculations and correlated methods (SCS-MP2 and SCS-CC2) predict that Im·Bz has a Cs-symmetric T-shaped minimum-energy structure with an NH···π hydrogen bond to the Bz ring; the NH bond is tilted 12° away from the Bz C6 axis. IR depletion spectra support the T-shaped geometry: The NH stretch vibrational fundamental is red shifted by -73 cm(-1) relative to that of bare imidazole at 3518 cm(-1), indicating a moderately strong NH···π interaction. While the S0(A1g) → S1(B2u) origin of benzene at 38 086 cm(–1) is forbidden in the gas phase, Im·Bz exhibits a moderately intense S0 → S1 origin, which appears via the D(6h) → Cs symmetry lowering of Bz by its interaction with imidazole. The NH···π ground-state hydrogen bond is strong, De=22.7 kJ/mol (1899 cm–1). The combination of gas-phase UV and IR spectra confirms the theoretical predictions that the optimum Im·Bz geometry is T shaped and NH···π hydrogen bonded. We find no experimental evidence for a CH···π hydrogen-bonded ground-state isomer of Im·Bz. The optimum NH···π geometry of the Im·Bz complex is very different from the majority of the histidine·aromatic contact geometries found in protein database analyses, implying that the CH···π contacts observed in these searches do not arise from favorable binding interactions but merely from protein side-chain folding and crystal-packing constraints. The UV and IR spectra of the imidazole·(benzene)2 cluster are observed via fragmentation into the Im·Bz+ mass channel. The spectra of Im·Bz and Im·Bz2 are cleanly separable by IR hole burning. The UV spectrum of Im·Bz2 exhibits two 000 bands corresponding to the S0 → S1 excitations of the two inequivalent benzenes, which are symmetrically shifted by -86/+88 cm(-1) relative to the 000 band of benzene

1147 related Products with: Modeling the Histidine-Phenylalanine Interaction: The NH···π Hydrogen Bond of Imidazole·Benzene.

BACTERIOLOGY BACTEROIDES TCP-1 theta antibody Sour Recombinant Thermostable Recombinant Thermostable Recombinant Thermostable Recombinant Human PKC the Recombinant Human PKC the Recombinant Human PKC the Single Strand DNA Ligase, Single Strand DNA Ligase, Thermostable TDG Enzyme & Thermostable TDG Kit

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#25795666   2015/05/06 Save this To Up

Fermentation and Cost-Effective 13C/15N Labeling of the Nonribosomal Peptide Gramicidin S for Nuclear Magnetic Resonance Structure Analysis.

Gramicidin S (GS) is a nonribosomally synthesized decapeptide from Aneurinibacillus migulanus. Its pronounced antibiotic activity is attributed to amphiphilic structure and enables GS interaction with bacterial membranes. Despite its medical use for over 70 years, the peptide-lipid interactions of GS and its molecular mechanism of action are still not fully understood. Therefore, a comprehensive structural analysis of isotope-labeled GS needs to be performed in its biologically relevant membrane-bound state, using advanced solid-state nuclear magnetic resonance (NMR) spectroscopy. Here, we describe an efficient method for producing the uniformly (13)C/(15)N-labeled peptide in a minimal medium supplemented by selected amino acids. As GS is an intracellular product of A. migulanus, we characterized the producer strain DSM 5759 (rough-convex phenotype) and examined its biosynthetic activity in terms of absolute and biomass-dependent peptide accumulation. We found that the addition of either arginine or ornithine increases the yield only at very high supplementing concentrations (1% and 0.4%, respectively) of these expensive (13)C/(15)N-labeled amino acids. The most cost-effective production of (13)C/(15)N-GS, giving up to 90 mg per gram of dry cell weight, was achieved in a minimal medium containing 1% (13)C-glycerol and 0.5% (15)N-ammonium sulfate, supplemented with only 0.025% of (13)C/(15)N-phenylalanine. The 100% efficiency of labeling is corroborated by mass spectrometry and preliminary solid-state NMR structure analysis of the labeled peptide in the membrane-bound state.

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5(6) FAM, SE [5 (and 6) C 5(6) FAM, SE [5 (and 6) C 5 FAM, SE [5 Carboxyfluor 5 FAM, SE [5 Carboxyfluor 5(6) TAMRA, SE [5 (and 6) 5(6) TAMRA, SE [5 (and 6) 5 TAMRA, SE [5 Carboxytet 5 TAMRA, SE [5 Carboxytet CAR,Car,Constitutive andr CAR,CAR,Constitutive acti CAR,Car,Constitutive andr 4 ((4 (dimethylamino)phen

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#24992776   2014/07/04 Save this To Up

Amniotic fluid amino acid concentrations in fetal skeletal dysplasia.

The authors' objective was to measure amniotic fluid amino acid concentrations in pregnant women diagnosed as having fetuses with skeletal dysplasia in the second trimester of pregnancy. Eighteen pregnant women who had fetuses with with skeletal dysplasia detected by ultrasonography (skeletal dysplasia) in the second trimester and 35 women who had abnormal triple screenings indicating an increased risk for Down syndrome, but had healthy fetuses (control group), were enrolled in the study. Amniotic fluid was obtained by amniocentesis. The chromosomal analysis of the study and control groups was normal. Levels of free amino acids and non-essential amino acids were measured in amniotic fluid samples using GC/FID free (physiological) amino acid kit by gas chromatography. The mean levels of essential amino acids (histidine, isoleucine, leucine, lysine, methionine, phenylalanine, threonine, tryptophan, and valine) in amniotic fluid were found to be significantly lower in fetuses with skeletal dysplasia than in the control group (p < 0.05). The detection of significantly lower amino acid concentrations in the amniotic fluid of fetuses with a skeletal dysplasia compared to healthy fetuses suggests amino acid deficiency may play an etiological role in the pathogenesis of skeletal dysplasia.

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#24842400   2014/07/01 Save this To Up

Quantification of recombinant human erythropoietin by amino acid analysis using isotope dilution liquid chromatography-tandem mass spectrometry.

Herein, we describe an accurate method for protein quantification based on conventional acid hydrolysis and an isotope dilution-ultra performance liquid chromatography-tandem mass spectrometry method. The analyte protein, recombinant human erythropoietin (rhEPO), was effectively hydrolyzed by incubation with 8 mol/L hydrochloric acid at 130 °C for 48 h, in which at least 1 μmol/kg of rhEPO was treated to avoid possible degradation of released amino acids during hydrolysis. Prior to hydrolysis, sample solution was subjected to ultrafiltration to eliminate potential interfering substances. In a reversed-phase column, the analytes (phenylalanine, proline, and valine) were separated within 3 min using gradient elution comprising 20 % (v/v) acetonitrile and 10 mmol/L ammonium acetate, both containing 0.3 % (v/v) trifluoroacetic acid. The optimized hydrolysis and analytical conditions in our study were strictly validated in terms of accuracy and precision, and were suitable for the accurate quantification of rhEPO. Certified rhEPO was analyzed using a conventional biochemical assay kit as an additional working calibrant for the quantification of EPO and improved the accuracy. The optimized protocol is suitable for the accurate quantification of rhEPO and satisfactorily serves as a reference analytical procedure for the certification of rhEPO and similar proteins.

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#23812855   2013/09/19 Save this To Up

Enantioseparation of 6-aminoquinolyl-N-hydroxysuccinimidyl carbamate tagged amino acids and other zwitterionic compounds on cinchona-based chiral stationary phases.

The fluorescent tag 6-aminoquinolyl-N-hydroxysuccinimidyl carbamate (AQC; AccQ Fluor reagent kit from Waters) is a commercial N-terminal label for proteinogenic amino acids (AAs), designed for reversed-phase separation and quantification of the AA racemates. The applicability of AQC-tagged AAs and AA-type zwitterionic compounds was tested for enantiomer separation on the tert-butyl carbamate modified quinine and quinidine based chiral stationary phases, QN-AX and QD-AX employing polar-organic elution conditions. The investigated test analytes included the enantiomers of the positional isomers of isoleucine (Ile), threonine, homoserine, and 4-hydroxyproline. Furthermore, β-AAs, cyclic, and heterocyclic AAs including trans-2-amino-cyclohexane carboxylic acid and trans-2-aminocyclohexyl sulfonic acid, phenylalanine derivatives substituted with halides with increasing electronegativity and 3,4-dihydroxyphenylalanine, cysteine-related derivatives including homocysteic acid, methionine sulfone, cysteine-S-acetic acid, and cysteine-S-acetamide as well as a small range of aminophosphonic acids were enantioseparated. A mechanistic interaction study of AQC-AAs in comparison with fluoresceine isothiocyanate-labeled AAs was performed. The chiral and chemoselective recognition processes involved in enantiomer separation and retention was systematically discussed. Special emphasis was set on the influential factors exhibited by the chemistry, branching position, and spatial properties of the investigated zwitterionic analytes. The general interest to separate and distinguish between different types of branched-chained AAs and metabolic side products thereof lies in the toxicity of some of these compounds, which makes for instance allo-Ile an attractive candidate in disease-related biomarker research.

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6-Aminoquinolyl-N-hydroxy Fibroblast Growth Factor Fibroblast Growth Factor anti GFP antibody, rat mo (E)-O-(2-Acetamido-2-deox (Z)-O-(2-Acetamido-2-deox O-(2-Acetamido-2-deoxy-3, O-(2-Acetamido-3,4,6-tri- (1R,2R,3R)-(4S)-Amino-1,2 Benzyl [2-Amino-2-(hydrox Benzyl [1-[4-[[(4-Fluorob Benzyl [1-[4-[[(4-Fluorob

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#23673571   2013/09/19 Save this To Up

Amino acid analysis of dried blood spots for diagnosis of phenylketonuria using capillary electrophoresis-mass spectrometry equipped with a sheathless electrospray ionization interface.

We describe a capillary electrophoresis-mass spectrometry (CE-MS) method for newborn screening of a representative amino acid metabolic disease, namely, phenylketonuria (PKU). Underivatized phenylalanine and tyrosine in a dried blood spot (DBS) were simultaneously determined by CE-MS equipped with an ionophore membrane-packed sheathless electrospray ionization interface, which was developed by our group. The method was optimized for rapid determination of the underivatized amino acids, phenylalanine and tyrosine extracted from a DBS. Under the optimized conditions, the limit of detection of phenylalanine and tyrosine (signal-to-noise ratio, 3) was 0.03 and 0.07 mg/L in DBS, respectively, with a CE run time of less than 3 min. For repeated runs of a sample, coefficients of variation (CVs) for migration time were less than 3.7%, whereas CVs for the area ratio under the curve were 2.1 and 2.9% for 20 consecutive runs of 49.5 mg/kg Phe and 36.2 mg/kg Tyr, respectively. However, the relative standard deviations of intra- and interday assays for DBS samples were <6.2 and <5.8%, respectively, which were substantially due to sample extraction from DBS. The analytical method was applied to real clinical samples of Korean neonates, and results were compared with those of conventional methods for PKU diagnosis, which required reference analytical methods such as isotope dilution CE-MS or high-performance liquid chromatography-mass spectrometry for quality assurance of the conventional kit-based assays. The distinct advantages of high sensitivity and extremely low sample volume, as well as a simple, easy, and economic sample pretreatment, were demonstrated for the proposed method.

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Fibroblast Growth Factor Fibroblast Growth Factor DNP X acid [6 (2,4 Dinitr DNP X acid, SE [6 (2,4 Di Primary antibody GFR alp anti GFP antibody, rat mo EDANS acid [5 ((2 Aminoet EDANS acid [5 ((2 Aminoet EDANS sodium salt [5 ((2 EDANS iodoacetamide [5 (( N (2 Acetamido) 2 aminoet 2-Acetamido-5-aminophenyl

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