gebhart/patrepeval huggingface dataset:
Mutational Analysis, Using a Full-Length Rubella Virus cDNA Clone, of Rubella Virus E1 Transmembrane and Cytoplasmic Domains Required for Virus Release (1999)

We report on the construction of a full-length cDNA clone, pBRM33, derived from wild-type rubella virus M33 strain. The RNA transcripts synthesized in vitro from pBRM33 are highly infectious, and the viruses produced retain the phenotypic characteristics of the parental M33 virus in growth rate and plaque size. This cDNA clone was used to study the role of E1 transmembrane and cytoplasmic domains in virus assembly by site-directed mutagenesis. Three different alanine substitutions were introduced in the transmembrane domain of E1. These included substitution of leucine 464, cysteine 466, cysteine 467, and both cysteines 466 and 467 to alanine. In the E1 cytoplasmic domain, cysteine 470 and leucine 471 were altered to alanine. We found that these mutations did not significantly affect viral RNA replication, viral structural protein synthesis and transport, or E2/E1 heterodimer formation. Except for the substitution of cysteine 470, these mutations did, however, lead to a reduction in virus release. Substitution of cysteine 467 in the transmembrane region and of leucine 471 in the cytoplasmic domain dramatically reduced virus yield, resulting in the production of only 1 and 10% of the parental virus yield, respectively, in a parallel infection. These data show that E1 transmembrane and cytoplasmic domains play an important role in late stages of virus assembly, possibly during virus budding, consistent with earlier studies indicating that the E1 cytoplasmic domain may interact with nucleocapsids and that this interaction drives virus budding.
Antitumor Agents. 168. Dysoxylum cumingianum. IV. The Structures of Cumingianosides G-O, New Triterpene Glucosides with a 14,18-Cycloapotirucallane-Type Skeleton from Dysoxylum cumingianum, and Their Cytotoxicity against Human Cancer Cell Lines. (1997)

Nine new triterpene glucosides, named cumingianosides G-O (4-7, 9, 12-15), containing a 14,18-cycloapotirucallane-type skeleton were isolated from a cytotoxic fraction of the leaves of Dysoxylum cumingianum. The structures of the new compounds were established on the basis of chemical and spectral examinations. Evaluation of the cytotoxic activity of cumingianosides G-O showed that cumingianoside M exhibited significant (< 4 microM) cytotoxicity, especially against leukemia and melanoma cell lines.
Molecular Pharmacology: A Short Course (1997)

1. Biologic Systems and Definitions of Terms 2. Dose-Response Curves 3. Molecular Models of Drug-Receptor Interaction 4. Human Receptors from Recombinant DNA 5. Concentrations of Drugs in Tissues 6. Agonism and Stimulus-Response Mechanisms 7. Drug-Response Measurement Systems: Signal Strength, Stability, and Selectivity 8. Drug Antagonism 9. Statistical and Biologic Significance
Method to express and purify nm23-H2 protein from baculovirus-infected cells (2003)

High-throughput protein expression and purification are major bottlenecks in the postgenomic and proteomic era. We show here an automated method to express and purify nm23-H2, a nucleoside diphosphate kinase (NDPK), in a 96-well format, by the use of a robotic workstation, from insect Spodoptera frugiperda (Sf9) baculovirus-infected cells using nickel-nitrilotriacetic acid (Ni-NTA) agarose beads. The automated method is coupled to mass spectrometry for a validation and quality-control analysis. To verify the bona fide of the recombinant protein, several tests have been produced, including NDPK assay, Western blotting, and in vitro phosphorylation experiments, thus confirming the value of the protocol developed. The method has been validated for the expression of several proteins, thus confirming the value of this automated protocol. The research presented here is a useful method both for industrial and academic environments to produce in a high-throughput mode recombinant eukaryotic proteins to be assayed for a specific function in a systematic manner.
Distributed measurement of static strain in an optical fiber with multiple Bragg gratings at nominally equal wavelengths (1998)

The Fourier transform relationship between the reflected light froma Bragg grating and the complex spatial modulation of the Bragg grating is used to produce a distributed strain sensing system. A tunable external cavity diode laser along with a reference reflector in anoptical fiber are used to produce a measurement of the phase and amplitude of the reflected light from the modulated Bragg grating as a function of wavelength. The system is demonstrated with 22 Bragg gratings in a single fiber on a cantilever beam and compared with foil strain gauge readings.
Identification of â€œsafe harborâ€ loci in indica rice genome by harnessing the property of zinc-finger nucleases to induce DNA damage and repair (2014)

Zinc-finger nucleases (ZFNs) have proved to be successful tools for targeted genome manipulation in several organisms. Their main property is the induction of double-strand breaks (DSBs) at specific sites, which are further repaired through homologous recombination (HR) or non-homologous end joining (NHEJ). However, for the appropriate integration of genes at specific chromosomal locations, proper sites for gene integration need to be identified. These regions, hereby named safe harbor loci, must be localized in non-coding regions and possess high gene expression. In the present study, three different ZFN constructs (pZFN1, pZFN2, pZFN3), harboring β-glucuronidase (GUS) as a reporter gene, were used to identify safe harbor loci on rice chromosomes. The constructs were delivered into IR64 rice by using an improved Agrobacterium-mediated transformation protocol, based on the use of immature embryos. Gene expression was measured by histochemical GUS activity and the flanking regions were determined through thermal-asymmetric interlaced polymerase chain reaction (TAIL PCR). Following sequencing, 28 regions were identified as putative sites for safe integration, but only one was localized in a non-coding region and also possessed high GUS expression. These findings have significant applicability to create crops with new and valuable traits, since the site can be subsequently used to stably introduce one or more genes in a targeted manner.
Astatine-211 labeling of an antimelanoma antibody and its Fab fragment using N-succinimidyl p-[211At]astatobenzoate: comparisons in vivo with the p-[125I]iodobenzoyl conjugate (1991)

Astatine-211 labeling of an antimelanoma antibody, NR-ML-05, and its Fab fragment with N-succinimidyl p-[211At]astatobenzoate (2a) has been described. Preparation of the astatinated intermediate 2a was accomplished by distilling astatine-211 from an irradiated bismuth target directly into a reaction mixture containing an organometallic compound, N-succinimidyl p-(tri-n-butylstannyl)benzoate (1), and an oxidant, N-chlorosuccinimide, in 5% HOAc/MeOH. Trapping of distilled astatine as 2a was found to be efficient, resulting in 70-90% yields based on the amount of astatine-211 in the reaction mixture. The dry distillation technique employed gave recoveries of astatine-211 which ranged from 20% to 75%. Conjugation of 2a to NR-ML-05 and its Fab fragment was accomplished in 40-60% yields. The [211At]astatobenzoyl-conjugated antibodies were found to be stable in vitro when challenged by strong denaturants and nucleophilic reagents. Coinjected dual-labeled studies of the 2a astatinated antibodies and the same antibodies labeled with N-succinimidyl p-[125I]iodobenzoate (2b) in athymic mice bearing the human tumor xenograft A375 Met/Mix demonstrated that both radiolabeled antibodies had equivalent tumor localization. Data from the dual-labeled biodistribution of the intact antibody suggests that the astatine is stably attached. Data from the dual-labeled Fab fragment suggests that a portion of the astatine label is released as astatide, either from the astatinated Fab or from a catabolite.
Dapsone-Induced Hemolytic Anemia (1995)

Dapsone, an old drug introduced and used almost exclusively for the treatment of leprosy, is now utilized in an increasing number of therapeutic situations. However, its hemotoxicity is potentially severe and is often dose limiting. Effective countermeasures, based on resolution of the mechanisms underlying dapsone-induced hemotoxicity, could significantly enhance the therapeutic value of the drug. In studies on rat red cells, we have established that the N-hydroxy metabolites of dapsone, DDS-NOH and MADDS-NOH, are direct-acting hemolytic agents, that they are formed in amounts sufficient to account for the hemotoxicity of the parent drug, and that the action of these toxic metabolites in the red cell induces premature sequestration by the spleen. Incubation of rat red cells with hemolytic concentrations of arylhydroxylamines leads to the generation of hydroxyl, glutathiyl, and hemoglobinthiyl radicals, and the formation of protein-glutathione mixed disulfides. Disulfide-linked adducts are also formed between membrane skeletal proteins and hemoglobin monomers, as well as between the monomeric hemoglobin units forming dimers, trimers, tetramers, and pentamers. Profound morphological changes are seen with change from normal discoidocity to an extreme nonspherocytic enchinocyte shape. Parallel studies with human red cells indicate that the response of human cells is qualitatively similar but that there are notable differences in regard to skeletal membrane effects. A working hypothesis for the mechanism underlying dapsone hemolytic activity is proposed.
Analysis on the Impact of Fracturing Treatment Design and Reservoir Properties on Production from Shale Gas Reservoirs (2013)

Abstract Production from shale gas reservoirs depends greatly on the efficiency of hydraulic fracturing treatments. The cumulated experience in the industry has led to several best practices in treatment design, which have improved productivity of these reservoirs. However, further advancement of treatment design requires a deeper understanding of the complex physics involved in both hydraulic fracturing and production, such as stress shadow, proppant placement and treatment interaction with pre-existing natural fractures. This paper sheds light on the non-linear physics involved in the production of shale gas reservoirs by improving the understanding of the complex relation between gas production, the reservoir properties, and several treatment design parameters. A fracturing-to-production simulation workflow integrating the Unconventional Fracture Model (Weng et al., 2011), with the Unconventional Production Model (Cohen et al., 2012) is presented. By applying this workflow to a realistic reservoir, we did an extensive parametric study to investigate the relation between production and treatment design parameters such as fracturing fluid viscosity, proppant size, proppant concentration, proppant injection order, treatment volume, pumping rate, pad size and hybrid treatment. The paper also evaluates the influence of unconventional reservoir properties - such as permeability, horizontal stress, horizontal stress anisotropy, horizontal stress orientation, Poisson's ratio and Young's modulus - on production. Since this paper focuses on fluid and proppant selection, our methodology was to run 28 simulations to cover the 2D parametric space of proppant size and fracturing fluid viscosity for all of these parameters. More than fourteen hundred simulations were run in this parametric study and the results provide guidelines for optimized treatment design. This paper illustrates how this unique workflow can identifies the optimum fluid and proppant selection that gives the maximum production for a given reservoir and completion. In addition, the parametric study shows how these optimums evolve as a function of reservoir and treatment parameters. The results validate several best practices in treatment design for shale. For example, combination of different sizes of proppant optimizes production by maximizing initial production and slowing down production decline. Simulations also confirm the best practice of injecting the smallest proppant first. The study explains why slickwater treatments should be injected at maximum pumping rate and preferably with 40/70 mesh sand. It also illustrates why reservoirs with high Young's modulus (such as the Barnett shale) can be stimulated effectively with slickwater. Another key finding is that the optimum fluid viscosity increases with treatment volume.
Core technologies for service robotics (2005)

Service robotics products are becoming a reality. This paper describes three core technologies that enable the next generation of service robots. They are low-cost, make use of low-cost hardware, and prepare for a short time-to-market for product development. The first technology is an object recognition system, which can be used by the robot to interact with the environment The second technology is a vision-based navigation system (vSLAM/spl trade/), which simultaneously can build a map and localize the robot in the map. Finally, the third technology is a flexible and rich software platform (ERSP/spl trade/) that assists developers in rapid design and prototyping of robotics applications.
Lovastatin Lactone Inhibits Methane Production in Human Stool Homogenates (2015)

Introduction: Research has linked an altered gut microbiome to symptoms of irritable bowel syndrome (IBS). In constipation-predominant IBS (IBS-C), methane production appears important as methane levels on breath are proportional to the degree of constipation, and antibiotic elimination of methane significantly improves constipation. The predominant methane-producing organism in humans is Methanobrevibacter smithii. Recent data suggests certain HMG-CoA reductase inhibitors (statins) may reduce methane production by inhibiting methanogenesis. This study examined the effects of various statins on methane production in fresh human stool homogenates. Methods: Five female subjects recruited based on high breath methane production (average=69.6ppm) provided a total of 8 fresh stool samples each. Samples were homogenized in 1XPBS (2g stool/3ml PBS) in an anaerobic chamber at 37°C and divided into 5 stoppered flasks. To assess statin effects, head gas withdrawn through a stopcock was collected at baseline, then every 30min for a total of 270min, and analyzed on a Quintron Model SC gas chromatograph for methane levels. Nine statins were initially assessed for methane inhibition at a concentration of 5mg/g stool: lovastatin lactone and hydroxyacid, pravastatin lactone and hydroxyacid, simvastatin lactone, mevastatin lactone, rosuvastatin lactone, and atorvastatin lactone and hydroxyacid. Once the ideal statin was determined, the effects of concentrations of 0.04, 0.12, 0.48, 1, 5, and 10mg/g stool were tested by sampling head gas every 90min after statin addition, up to 720min.Further assessments were then performed comparing three forms of lovastatin (lactone, diol and hydroxyacid). Results: Lovastatin lactone was identified as the only effective methane inhibitor (Figure 1), significantly inhibiting methane levels by -65% of the control stool. Lovastatin lactone at 5mg/g produced the maximum inhibiting effect resulting in an average methane level of 3% of the control over time (Figure 2). In a final validation comparison of 3 forms of lovastatin (5mg/g), both the lactone and diol forms proved to be effective. In all assessments, hydroxyacid forms were least able to inhibit methane production in fresh stool samples.Figure 1Figure 2Conclusion: Lovastatin lactone and lovastatin diol at concentrations of 5mg/g stool significantly reduce methane production in human stool homogenates. Future studies are needed to assess the effect of this statin on methane production and IBS-C symptoms in humans.
Serum-free Erythroid Differentiation for Efficient Genetic Modification and High-Level Adult Hemoglobin Production (2018)

<i>In vitro</i> erythroid differentiation from primary human cells is valuable to develop genetic strategies for hemoglobin disorders. However, current erythroid differentiation methods are encumbered by modest transduction rates and high baseline fetal hemoglobin production. In this study, we sought to improve both genetic modification and hemoglobin production among human erythroid cells <i>in vitro</i>. To model therapeutic strategies, we transduced human CD34<sup>+</sup> cells and peripheral blood mononuclear cells (PBMCs) with lentiviral vectors and compared erythropoietin-based erythroid differentiation using fetal-bovine-serum-containing media and serum-free media. We observed more efficient transduction (85%-93%) in serum-free media than serum-containing media (20%-69%), whereas the addition of knockout serum replacement (KSR) was required for serum-free media to promote efficient erythroid differentiation (96%). High-level adult hemoglobin production detectable by electrophoresis was achieved using serum-free media similar to serum-containing media. Importantly, low fetal hemoglobin production was observed in the optimized serum-free media. Using KSR-containing, serum-free erythroid differentiation media, therapeutic adult hemoglobin production was detected at protein levels with β-globin lentiviral transduction in both CD34<sup>+</sup> cells and PBMCs from sickle cell disease subjects. Our <i>in vitro</i> erythroid differentiation system provides a practical evaluation platform for adult hemoglobin production among human erythroid cells following genetic manipulation.
Preparation and hybridization properties of oligonucleotides containing 1-α-D-arabinofuranosylthymine (1991)

A pentadecanucleotide was prepared from 1-alpha-arabinofuranosylthymine. This novel oligonucleotide was found to hybridize to oligodeoxyadenylate, although not a s strongly as pentadecathymidylate. It formed duplex hybrids with both DNA and RNA complements, and triplex structures with a duplex containing a (dT)15-(dA)15 tract within a more complex strand. The Tm of the duplex with polyadenylate was almost the same as that of (dT)15 and polyadenylate, while its Tm with (dA)15 was substantially lower than that of the natural counterpart. A selective benzoylation of the 2'-O of a 5'-blocked alpha-ara-thymine was developed to greatly simplify the preparation of suitable blocked material for use in preparation of oligonucleotides on the automated DNA synthesizer.
The generation of a humanized, non‐mitogenic CD3 monoclonal antibody which retains in vitro immunosuppressive properties (1993)

CD3 antibodies are proven immunosuppressants capable of reversing transplant rejection episodes. Their general application has been limited both by their immunogenicity and, in particular, by the "first-dose" cytokine-release syndrome experienced by patients after the initial administration of antibody. We have produced a set of variants of the humanized YTH 12.5 CD3 monoclonal antibody (mAb) (Routledge et al., Eur. J. Immunol. 1991. 21: 2717) bearing different human heavy (H) chain constant regions, with the intention of finding a form of the antibody that is not able to activate T cells. Comparison of the variants having gamma 1, gamma 2, gamma 3 and gamma 4 H chains in a competitive binding assay showed that antibody avidity was not affected by IgG subclass. Using a sensitive indicator of FcR binding activity (the capacity of the CD3 mAb to redirect cytotoxic T cells to kill the monocytic cell line U-937) we demonstrated a functional hierarchy of gamma 1 = gamma 4 > alpha 2 =/> gamma 3 mb >> gamma 2. An aglycosyl version of the gamma 1 CD3 mAb, produced by site-directed mutagenesis (Asn297 to Ala), still had considerable activity in this assay (intermediate to the gamma 1 and alpha 2 CD3 mAb), albeit at a level approximately 10-fold lower than that of the parental gamma 1 form. When we tested their ability to stimulate T cell proliferation in vitro in the presence of 5% human serum, all of the wild-type immunoglobulin isotypes were found to be active, although there were T cell donor-dependent variations in the extent of the responses. The aglycosyl gamma 1 mAb was, however, completely non-mitogenic in all of ten donors tested, unless the assay was performed in IgG-free medium. Despite being non-stimulatory, this mAb was also able to inhibit the mixed lymphocyte reaction responses of both naive and primed T cells. Comparison of the gamma 1 and aglycosyl gamma 1 mAb in an experimental mouse model for CD3 mAb-induced cytokine release indicated that removal of the carbohydrate moiety from the gamma 1 constant region reduced the in vivo tumor necrosis factor-alpha response by a factor of at least 16-fold. These data suggest that the aglycosyl gamma 1 CD3 mAb is a promising candidate for immunosuppressive therapy without "first dose" side effects.
An active site homology model of phenylalanine ammonia‐lyase from P. crispum (2002)

The plant enzyme phenylalanine ammonia-lyase (PAL, EC 4.3.1.5) shows homology to histidine ammonia-lyase (HAL) whose structure has been solved by X-ray crystallography. Based on amino-acid sequence alignment of the two enzymes, mutagenesis was performed on amino-acid residues that were identical or similar to the active site residues in HAL to gain insight into the importance of this residues in PAL for substrate binding or catalysis. We mutated the following amino-acid residues: S203, R354, Y110, Y351, N260, Q348, F400, Q488 and L138. Determination of the kinetic constants of the overexpressed and purified enzymes revealed that mutagenesis led in each case to diminished activity. Mutants S203A, R354A and Y351F showed a decrease in kcat by factors of 435, 130 and 235, respectively. Mutants F400A, Q488A and L138H showed a 345-, 615- and 14-fold lower kcat, respectively. The greatest loss of activity occurred in the PAL mutants N260A, Q348A and Y110F, which were 2700, 2370 and 75 000 times less active than wild-type PAL. To elucidate the possible function of the mutated amino-acid residues in PAL we built a homology model of PAL based on structural data of HAL and mutagenesis experiments with PAL. The homology model of PAL showed that the active site of PAL resembles the active site of HAL. This allowed us to propose possible roles for the corresponding residues in PAL catalysis.
Two-site substrate recognition model for the Keap1-Nrf2 system: a hinge and latch mechanism (2006)

Cells are equipped with a number of transcriptional factors that safeguard against various environmental insults. Proteasomal protein degradation plays an important role in the Keap1-Nrf2 cytoprotection system, with molecular machinery similar to that for other environmental defense systems such as inflammatory and hypoxic responses. While Nrf2 protein stabilization is known to be redox-sensitive, the transcription factors NF-kappaB and HIF-1alpha for inflammatory and hypoxic responses, respectively, are also influenced by the cellular redox conditions. In this review we present the recently proposed two-site substrate recognition model of the Keap1-Nrf2 system, which regulates the cellular responses against oxidative and xenobiotic stresses. The implications of two destructive motifs in Nrf2, the ETGE and DLG motifs, which appear to function as a hinge and latch attenuating Keap1 activity in different redox states, are discussed.
Ascorbate-Nitrite Reaction: Possible Means of Blocking the Formation of Carcinogenic N -Nitroso Compounds (1972)

The formation of carcinogenic N-nitroso compounds by the chemical reaction between nitrous acid and oxytetracycline, morpholine, piperazine, N-methylaniline, methylurea, and (in some experiments) dimethylamine was blocked by ascorbic acid. The extent of blocking depended on the compound nitrosated and on the experimental conditions. Urea and ammonium sulfamate were less effective as blocking agents. The possibility of in vivo formation of carcinogenic N-nitroso compounds from drugs could be lessened by the combination of such drugs with the ascorbic acid.