gebhart/patrepeval huggingface dataset:
A Highly Effective Dominant Negative αs Construct Containing Mutations That Affect Distinct Functions Inhibits Multiple Gs-coupled Receptor Signaling Pathways (2002)

To investigate the subcellular organization of receptor-G protein signaling pathways, a robust dominant negative alpha(s) mutant containing substitutions that alter distinct functions was produced and tested for its effects on G(s)-coupled receptor activity in HEK-293 cells. Mutations in the alpha3beta5 loop region, which increase receptor affinity, decrease receptor-mediated activation, and impair activation of adenylyl cyclase, were combined with G226A, which increases affinity for betagamma, and A366S, which decreases affinity for GDP. This triple alpha(s) mutant can inhibit signaling to G(s) from the luteinizing hormone receptor by 97% and from the calcitonin receptor by 100%. In addition, this alpha(s) mutant blocks all signaling from the calcitonin receptor to G(q). These results lead to two conclusions about receptor-G protein signaling. First, individual receptors have access to multiple types of G proteins in HEK-293 cell membranes. Second, different G protein alpha subunits can compete with each other for binding to the same receptor. This dominant negative alpha(s) construct will be useful for determining interrelationships among distinct receptor-G protein interactions in a wide variety of cells and tissues.
Viral Agents Oncolytic for Human Tumors in Heterologous Host. Oncolytic Effect of Coxsackie B Viruses. (1957)

Summary1. The Toolan technic for growing cancer tissue in a heterologous host was applied to growth of tissue culture propagated HeLa and KB cells in the peritoneal cavity of rat. This method resulted in rapid development of large healthy explants of carcinoma on peritoneal surfaces, particularly of omentum. Ninety-three% of rats inoculated developed comparatively uniform tumor masses, thus allowing for adequately controlled studies of virus growth and oncolytic effects. 2. A preliminary survey of adenovirus, ECHO, and Coxsackie agents in rat-HeLa tumor system produced 3 types of reaction: 1. failure of some agents to grow. 2. growth of virus without oncolytic activity, and, 3. growth of virus with evidence of slight or incomplete oncolytic effects. Four Coxsackie agents, types B1, B2, B3, and B5, showed initial evidence of growth, and on serial passage through rat-HeLa tumors, increased capacity for destroying solid carcinoimata in vivo, producing characteristic histological changes. The oncolytic Coxsackie B3 virus produced by rat-HeLa tumor passage in vivo, compared to the same strain passed in HeLa cells grown in tissue culture, revealed marked differences in oncolytic capacity. Use of this method illustrates the possibility of virus adaptation by tumor to tumor passage; that this principle has been successfully employed to destroy solid tumor transplants of human origin, suggests its possible application for future clinical studies.
Fidelity of DNA ligation: a novel experimental approach based on the polymerisation of libraries of oligonucleotides (1998)

Complete libraries of oligonucleotides were used as substrates for Thermus thermophilus DNA ligase, on a M13mp18 ssDNA template. A 17mer primer was used to start a polymerisation process. Ladders of ligation products were analysed by gel electrophoresis. Octa-, nona- and decanucleotide libraries were compared. Nonanucleotides were optimum for polymerisation and up to 15 monomers were ligated. The fidelity of incorporation was studied by sequencing 28 clones (2268 bases) of nonanucleotide polymers, 12 monomers in length. Of the ligated monomers, 79% were the correct complementary sequence. In a total of 57 (2.5%) mispaired bases, there was a strong bias to G.T, G.A, G.G and A.G mismatches. Of the mismatches, 86% were found to be purines on the incoming oligonucleotide, of which 71% were G. There is evidence for clustering of mismatches within specific 9mers and at specific positions within these 9mers. The most frequent mismatches were at the 5'-terminus of the oligonucleotide, followed by the central position. We suggest that sequence selection was imposed by the ligase and not just by base pairing interactions. The ligase directs polymerisation in the 3' to 5' direction which we propose is linked to its role in lagging strand DNA replication.
The Low Density Lipoprotein Receptor-related Protein Can Function Independently from Heparan Sulfate Proteoglycans in Tissue Factor Pathway Inhibitor Endocytosis (1996)

Tissue factor pathway inhibitor (TFPI) is a plasma serine protease inhibitor that directly inhibits coagulation factor Xa and regulates blood coagulation via inhibition of factor VIIa-tissue factor enzymatic activity. We previously demonstrated that >90% of TFPI bound to a single population of low affinity binding sites on hepatoma cells (2 x 10(6) sites/cell, Kd = 30 nM), and, that following binding, the low density lipoprotein receptor-related protein (LRP) mediated TFPI uptake and degradation. We subsequently reported heparan sulfate proteoglycans (HSPGs) constitute a second receptor system involved in TFPI catabolism. In the present study, mouse embryonic fibroblasts heterozygous and homozygous-negative for disruption of the LRP gene were used to further examine the roles of LRP and HSPGs in TFPI endocytosis. We demonstrate that LRP is absolutely required for degrading 125I-TFPI. LRP heterozygous and homozygous-negative cells bind 125I-TFPI similarly, and the 39-kDa protein, an inhibitor of all known ligand interactions with LRP, does not alter 125I-TFPI binding to these cells. TFPI can be cross-linked to LRP on [35S]cysteine-labeled hepatoma and LRP-heterozygous cells but not LRP-negative cells. When HSPGs are blocked with protamine, 125I-TFPI binds in a 39-kDa protein-inhibitable manner to 41,000 high affinity sites/hepatoma cell (Kd = 2.3 nM). Blockade of HSPGs with protamine results in significantly more 125I-TFPI degradation by LRP-positive cells. TFPI can be cross-linked to LRP in the absence and presence of protamine. However, in the presence of protamine, relative to the total pool of cross-linked proteins, 5-fold more TFPI is cross-linked to LRP. Finally, we show TFPI inhibits 125I-alpha2-macroglobulin-methylamine binding to hepatoma cells and that carboxyl-terminal residues 115-319 of the 39-kDa protein inhibit both 125I-TFPI degradation and binding when binding conditions contain protamine. Together, our results suggest that while the majority of TFPI binds to cell surface HSPGs, LRP can function independently from HSPGs in the binding and uptake of TFPI.
Expression and Role of VEGF-A in the Ciliary Body (2012)

VEGF-A has an important role in ciliary body homeostasis. Potential for undesired off-target effects should be considered with the chronic use of anti-VEGF-A therapies.
Artificially evolved Synechococcus PCC6301 Rubisco variants exhibit improvements in folding and catalytic efficiency (2007)

The photosynthetic CO2-fixing enzyme, Rubisco (ribulose-1,5-bisphosphate carboxylase/oxygenase), is responsible for most of the world's biomass, but is a slow non-specific catalyst. We seek to identify and overcome the chemical and biological constraints that limit the evolutionary potential of Rubisco in Nature. Recently, the horizontal transfer of Calvin cycle genes (rbcL, rbcS and prkA) from cyanobacteria (Synechococcus PCC6301) to gamma-proteobacteria (Escherichia coli) was emulated in the laboratory. Three unique Rubisco variants containing single (M259T) and double (M259T/A8S, M259T/F342S) amino acid substitutions in the L (large) subunit were identified after three rounds of random mutagenesis and selection in E. coli. Here we show that the M259T mutation did not increase steady-state levels of rbcL mRNA or L protein. It instead improved the yield of properly folded L subunit in E. coli 4-9-fold by decreasing its natural propensity to misfold in vivo and/or by enhancing its interaction with the GroES-GroEL chaperonins. The addition of osmolites to the growth media enhanced productive folding of the M259T L subunit relative to the wild-type L subunit, while overexpression of the trigger factor and DnaK/DnaJ/GrpE chaperones impeded Rubisco assembly. The evolved enzymes showed improvement in their kinetic properties with the M259T variant showing a 12% increase in carboxylation turnover rate (k(c)cat), a 15% improvement in its K(M) for CO2 and no change in its K(M) for ribulose-1,5-bisphosphate or its CO2/O2 selectivity. The results of the present study show that the directed evolution of the Synechococcus Rubisco in E. coli can elicit improvements in folding and catalytic efficiency.
Proto-oncogene c-kit expression in malignant melanoma: protein loss with tumor progression. (1997)

The c-kit gene encodes a transmembrane receptor that has tyrosine kinase activity. c-kit plays a role in hematopoiesis, gametogenesis, and melanogenesis. c-kit is found in melanocytes, and there is evidence that expression is lost in melanoma. We studied 85 melanocytic lesions for c-kit by immunohistochemical techniques using a monoclonal antibody. The lesions included banal nevi, junctional and compound nevi with melanocytic dysplasia, nontumorigenic radial growth phase melanoma, tumorigenic vertical growth phase melanoma, and metastatic melanoma. We found intense membrane staining in normal melanocytes and mast cells. Staining in compound nevi was strongest in junctional and superficial dermal components, whereas dermal nevi showed weak reactivity. Dysplastic nevi stained strongly, particularly in junctional cells. In melanoma, strong reactivity was most prominent in radial growth phase disease, but there was little or no staining in vertical growth phase and metastatic melanomas. In summary, c-kit protein is expressed in normal melanocytes, benign nevi, dysplastic nevi and nontumorigenic melanoma, but expression is lost in tumorigenic primary melanomas and metastases. The role of c-kit loss in advanced melanoma requires additional investigation.
Aspirin and alterations in DNA repair proteins in the SW480 colorectal cancer cell line (2010)

Regular aspirin intake is associated with a reduction in the incidence of colorectal cancer. Aspirin has been shown to be cytotoxic to colorectal cancer cells in vitro. The molecular basis for this cytotoxicity is controversial, with a number of competing hypotheses in circulation. One suggestion is that the protective effect is related to the induction of expression of the DNA mismatch repair (MMR) proteins hMLH1, hMSH2, hMSH6 and hPMS2 in DNA MMR proficient cells. We report that treatment of the DNA MMR competent/p53 mutant colorectal cancer cell line SW480 with 1 mM aspirin for 48 h caused changes in mRNA expression of several key genes involved in DNA damage signalling pathways, including a significant down-regulation in transcription of the genes ATR, BRCA1 and MAPK12. Increases in the transcription of XRCC3 and GADD45alpha genes are also reported. Regulation of these genes could potentially have profound effects on colorectal cancer cells and may play a role in the observed chemo-protective effect of aspirin in vivo. Although a correlation was not seen between transcript and protein levels of ATR, BRCA1 and GADD45alpha, an increase in XRCC3 encoded protein expression upon aspirin treatment in SW480 cells was observed by immunoblotting, immunofluorescence and immunohistochemical analysis. This is the first report of XRCC3 gene transcription and encoded protein expression being susceptible to exposure to the non-steroidal anti-inflammatory drug, aspirin. Furthermore, this study indicates that alterations in gene transcription seen in microarray studies must be verified at the protein level.
Deposition of Patterned Glycosaminoglycans on Silanized Glass Surfaces (2006)

We report the robust attachment of glycosaminoglycans (GAGs) on silanized glass surfaces. Depositions were performed both by immersion and by application of a pattern by means of microcontact printing. Immunofluorescence assays were performed to verify the deposition and the quality of the patterns. In addition, AFM studies of the coated surfaces were performed in order to study some physical characteristics of the deposited GAGs layers. These results may prove useful for the characterization of the mechanical properties of GAGs in the glycocalyx and its relation with cellular migration.
The I-TevI Nuclease and Linker Domains Contribute to the Specificity of Monomeric TALENs (2014)

Precise genome editing in complex genomes is enabled by engineered nucleases that can be programmed to cleave in a site-specific manner. Here, we fused the small, sequence-tolerant monomeric nuclease domain from the homing endonuclease I-TevI to transcription-like activator effectors (TALEs) to create monomeric Tev-TALE nucleases (Tev-mTALENs). Using the PthXo1 TALE scaffold to optimize the Tev-mTALEN architecture, we found that choice of the N-terminal fusion point on the TALE greatly influenced activity in yeast-based assays, and that the length of the linker used affected the optimal spacing of the TALE binding site from the I-TevI cleavage site, specified by the motif 5'-CNNNG-3'. By assaying activity on all 64 possible sequence variants of this motif, we discovered that in the Tev-mTALEN context, I-TevI prefers A/T-rich triplets over G/C-rich ones at the cleavage site. Profiling of nucleotide requirements in the DNA spacer that separates the CNNNG motif from the TALE binding site revealed substantial, but not complete, tolerance to sequence variation. Tev-mTALENs showed robust mutagenic activity on an episomal target in HEK 293T cells consistent with specific cleavage followed by nonhomologous end-joining repair. Our data substantiate the applicability of Tev-mTALENs as genome-editing tools but highlight DNA spacer and cleavage site nucleotide preferences that, while enhancing specificity, do confer moderate targeting constraints.
Inhibitors of mammalian G1 cyclin-dependent kinases. (1995)

C J Sherr and J M Roberts Howard Hughes Medical Institute, Department of Tumor Cell Biology, St. Jude Children's Research Hospital, Memphis, Tennessee 38104, USA.
Form and stability of aluminum hydroxide complexes in dilute solution (1967)

Structures of aqueous aluminum species_____________________________ Hydrolysis experiments.--
Coronary Artery Responses to Physiological Stimuli Are Improved by Deferoxamine but not by l -Arginine in Non–Insulin-Dependent Diabetic Patients With Angiographically Normal Coronary Arteries and No Other Risk Factors (1998)

This study shows that (1) responses of angiographically normal CAs to CPT and to flow increase are impaired in diabetic patients; (2) abnormal responses are not improved by L-arginine, suggesting that a deficit in substrate for nitric oxide synthesis is not involved; and (3) deferoxamine restores a vasodilator response to the two tests, suggesting that inactivation of NO by oxygen species might be partly responsible for the impairment of CA dilation in diabetic patients.
Estimation of DNA sequence divergence from comparison of restriction endonuclease digests (1977)

An estimation of the DNA sequence divergence between defined DNA secquences of individuals or species may be made from comparison by gel electrophoresis of restriction endonuclease digests. This analysis is applicable to purified DNA sequence of moderate complexity (1-100 X 10(6) daltons) which have diverged by base substitution of 0.5 to 25% of nucleotides.
Both ERK and Wnt/β-catenin pathways are involved in Wnt3a-induced proliferation (2004)

The Wnt family of proteins regulates development and cell growth. We identified Wnt3a-based regulatory mechanisms for cell proliferation in NIH3T3 fibroblast cells. The degree of Wnt3a-induced proliferation was reduced by beta-catenin small interfering RNA (siRNA) and extracellular signal-regulated kinase (ERK) siRNA, indicating that both the ERK and Wnt/beta-catenin pathways are involved in Wnt3a-induced proliferation. Wnt3a immediately and transiently activated the Raf-1-MEK-ERK cascade in a manner distinct from that of the beta-catenin increase seen in cells treated with Wnt3a. Wnt3a-induced ERK activation was maintained even though basal ERK activities were reduced by beta-catenin siRNA, indicating that Wnt3a may activate the ERK pathway independently of beta-catenin. The ERK pathway was however, activated by beta-catenin transfection, which was abolished by co-transfection with dominant-negative Tcf-4. Therefore, ERK pathway activation by Wnt signaling could occur at multiple levels, including beta-catenin-independent direct signaling resulting from a Wnt3a and beta-catenin/Tcf-4-dependent post gene transcriptional event. Wnt3a stimulated the G1 to S phase cell cycle progression. This stimulation was reduced by the ERK pathway inhibitor, indicating that Wnt3a promotes proliferation by stimulating the ERK pathway. Wnt3a therefore stimulates the proliferation of fibroblast cells, at least in part, via activation of the ERK and Wnt/beta-catenin pathways.
Overfitting and neural networks: conjugate gradient and backpropagation (2000)

Methods for controlling the bias/variance tradeoff typically assume that overfitting or overtraining is a global phenomenon. For multi-layer perceptron (MLP) neural networks, global parameters such as the training time, network size, or the amount of weight decay are commonly used to control the bias/variance tradeoff. However, the degree of overfitting can vary significantly throughout the input space of the model. We show that overselection of the degrees of freedom for an MLP trained with backpropagation can improve the approximation in regions of underfitting, while not significantly overfitting in other regions. This can be a significant advantage over other models. Furthermore, we show that "better" learning algorithms such as conjugate gradient can in fact lead to worse generalization, because they can be more prone to creating varying degrees of overfitting in different regions of the input space. While experimental results can not cover all practical situations, our results do help to explain common behavior that does not agree with theoretical expectations. Our results suggest one important reason for the relative success of MLPs, bring into question common beliefs about neural network training regarding training algorithms, overfitting, and optimal network size, suggest alternate guidelines for practical use, and help to direct future work.
Synthesis of Pyrimidine and Purine Bases. II (1965)

Several efforts were made on the simplification of the steps, the attainment of higher yields, and the reduction of material costs in the synthesis of adenine from malonic ester reported in a previous paper.(1) 4, 6-pyrimidinediol has been prepared in 80% yield from diethyl malonate and formamide by a one-step synthesis using ethanolic sodium ethoxide.(2) Nitration of 4, 6-pyrimidinediol could be carried out in 95% yield with a mixture of nitric acid and sulfuric acid.(3) Methanol could be used instead of ethanol as a solvent for amination of 4, 6-dichloro-5-nitropyrimidine to 4, 6-diamino-5-nitropyrimidine.With these improvements, adenine has been prepared in 35% over-all yield from diethyl malonate by a five-step synthesis.
Gut bacteria that prevent growth impairments transmitted by microbiota from malnourished children (2016)

Undernourished children exhibit impaired development of their gut microbiota. Transplanting microbiota from 6- and 18-month-old healthy or undernourished Malawian donors into young germ-free mice that were fed a Malawian diet revealed that immature microbiota from undernourished infants and children transmit impaired growth phenotypes. The representation of several age-discriminatory taxa in recipient animals correlated with lean body mass gain; liver, muscle, and brain metabolism; and bone morphology. Mice were cohoused shortly after receiving microbiota from healthy or severely stunted and underweight infants; age- and growth-discriminatory taxa from the microbiota of the former were able to invade that of the latter, which prevented growth impairments in recipient animals. Adding two invasive species, Ruminococcus gnavus and Clostridium symbiosum, to the microbiota from undernourished donors also ameliorated growth and metabolic abnormalities in recipient animals. These results provide evidence that microbiota immaturity is causally related to undernutrition and reveal potential therapeutic targets and agents.
Evidence by Mutagenesis that Tyr370 of the Mouse Ribonucleotide Reductase R2 Protein Is the Connecting Link in the Intersubunit Radical Transfer Pathway (1999)

Ribonucleotide reductase catalyzes all de novo synthesis of deoxyribonucleotides. The mammalian enzyme consists of two non-identical subunits, the R1 and R2 proteins, each inactive alone. The R1 subunit contains the active site, whereas the R2 protein harbors a binuclear iron center and a tyrosyl free radical essential for catalysis. It has been proposed that the radical properties of the R2 subunit are transferred approximately 35 A to the active site of the R1 protein, through a coupled electron/proton transfer along a conserved hydrogen-bonded chain, i.e. a radical transfer pathway (RTP). To gain a better insight into the properties and requirements of the proposed RTP, we have used site-directed mutagenesis to replace the conserved tyrosine 370 in the mouse R2 protein with tryptophan or phenylalanine. This residue is located close to the flexible C terminus, known to be essential for binding to the R1 protein. Our results strongly indicate that Tyr(370) links the RTP between the R1 and R2 proteins. Interruption of the hydrogen-bonded chain in Y370F inactivates the enzyme complex. Alteration of the same chain in Y370W slows down the RTP, resulting in a 58 times lower specific activity compared with the native R2 protein and a loss of the free radical during catalysis.
THE NEGLECTED SEPTAL CARTILAGE GRAFT: (WITH EXPERIMENTAL OBSERVATIONS ON THE GROWTH OF HUMAN CARTILAGE GRAFTS) (1945)

Many otolaryngologists and plastic surgeons appear to avoid the use of septal cartilage as a grafting material because of the mistaken impression that it is absorbed after transplantation. This erroneous belief induces the surgeon who is repairing a nose with an old depressed fracture to perform a submucous resection for the purpose of restoring breathing space but to discard the septal cartilage and bone which might be used as grafting material to fill the saddle depression. The patient is then subjected to additional surgical operation for removal of his rib cartilage, or else inferior material, such as preserved cadaveric cartilage, is used to restore the nasal bridge. In many of these cases the septal cartilage removed during the submucous resection would have provided sufficient grafting material for repair of the saddle. The general depreciation of septal cartilage as material for grafting is apparently based on derogatory statements which have been