bluuebunny/crossref_metadata_2025_split huggingface dataset:
Identification of different classes of superoxide dismutase in carnation petals
Authors: Droillard Marie‐Jo , Bureau Danielle , Paulin André , Daussant Jean 
Published: May 2005

AbstractIsoelectric focusing and nondenaturing two‐dimensional electrophoresis, followed by an enzymatic characterization involving use of specific inhibitors were applied to identify the different forms of the superoxide dismutase (SOD) extracted from petals of cut carnation. Electrophoresis was carried out either with the crude extract or with fractions obtained after ion exchange chromatography of the extract. Three forms of the SOD were identified (Cu, Zn‐SOD, Fe‐SOD and Mn‐SOD). They have close pI's (4.7 to 4.9) and most of them are composed of several isoforms.
Database and search techniques for two‐dimensional gel protein data: A comparison of paradigms for exploratory data analysis and prospects for biological modeling
Authors: Lemkin Peter F. , Lester Eric P. 
Published: May 2005

AbstractTwo‐dimensional (2‐D) polyacrylamide gel electrophoresis can detect thousands of polypeptides, separating them by apparent molecular weight (Mr) and isoelectric point (pI). Thus it provides a more realistic and global view of cellular genetic expression than any other technique. This technique has been useful for finding sets of key proteins of biological significance. However, a typical experiment with more than a few gels often results in an unwieldy data management problem. In this paper, the GELLAB‐II system is discussed with respect to how data reduction and exploratory data analysis can be aided by computer data management and statistical search techniques. By encoding the gel patterns in a “three‐dimensional” (3‐D) database, an exploratory data analysis can be carried out in an environment that might be called a “spread sheet for 2‐D gel protein data”. From such databases, complex parametric network models of protein expression during events such as differentiation might be constructed. For this, 2‐D gel databases must be able to include data from other domains external to the gel itself. Because of the increasing complexity of such databases, new tools are required to help manage this complexity. Two such tools, object‐oriented databases and expert‐system rule‐based analysis, are discussed in this context. Comparisons are made between GELLAB and other 2‐D gel databases analysis systems to illustrate some of the analysis paradigms common to these systems and where this technology may be heading.
Development of a database of amino acid sequences for proteins identified and isolated on two‐dimensional polyacrylamide gels
Authors: Sweatt J. David , Kennedy Timothy E. , Wager‐Smith Karen , Gawinowicz Mary A. , Barzilai Ari , Karl Kevin A. , Kandel Eric R. 
Published: May 2005

AbstractAs part of our continuing studies into the biochemical basis of long‐term changes in neuronal function in Aplysia, we have developed a simple method for obtaining amino acid sequence information from proteins isolated on two‐dimensional gels. Proteins isolated on preparative two‐dimensional gels are digested in situ with Staphylococcus, aureus V8 protease, and the resulting peptides electrophoresed, transferred to a polyvinylidene difluoride membrane, and sequenced in a gas‐phase sequencer. The method is simple and should be applicable to a variety of other systems where the development of a two‐dimensional gel database is underway.
A simple method for determination of the concentration of anti‐dextran IgG in antiserum by means of affinity electrophoresis
Authors: Tanaka Tatehiko , Nakamura Kazuyuki , Takeo Kazusuke 
Published: May 2005

AbstractA simple technique for determination of the concentration of anti‐dextran IgG in antiserum using affinity electrophoresis is described. In the presence of an excess of intermediary molecular size dextran in the polyacrylamide gel, polyclonal anti‐dextran IgG migrated in a single sharp band, separated from the nonspecific IgG fraction and other serum protein fractions. With this technique, 1–10 μg anti‐dextran IgG in antisera can be determined within 3 h. We call the procedure ligand saturating affinity electrophoresis.
Characterization of human residual catalase of an acatalasemic patient by isoelectric focusing and sodium dodecyl sulfate‐polyacrylamide gel electrophoresis followed by electrophoretic blotting and immunodetection
Authors: Ogata Masana , Suzuki Kazuhiko , Satoh Yukinori 
Published: May 2005

AbstractIsoelectric points and subunit sizes of catalases in human blood and human cultured skin fibroblasts from acatalasemic and normal subjects were analyzed by isoelectric focusing in agarose gel and by sodium dodecyl sulfate‐polyacrylamide gel electrophoresis, respectively, followed by electroblotting to polyvinylidene difluoride membranes for immunodetection. The results indicated that the isoelectric point of residual catalase in the C fraction prepared from acatalasemic erythrocytes was identical with that of catalase prepared from normal erythrocytes. The residual catalase in homogenates of acatalasemic cultured skin fibroblasts also reacted with anticatalase rabbit serum and had an isoelectric point identical with that of normal catalase. Sub‐unit sizes of normal and acatalasemic catalases in the C fractions of erythrocytes were also found to be identical on sodium dodecyl sulfate‐polyacrylamide gel electrophoresis, followed by electroblotting and immunoenzymatic amplification. The results indicated no substantial difference in molecular size and charge of catalase proteins between normal and acatalasemic erythrocytes and fibroblasts.
Use of a polynomial exponential function to describe migration of proteins on sodium dodecyl sulfate‐polyacrylamide gels
Authors: Winston Vern 
Published: May 2005

AbstractStandard mixtures of proteins were separated by sodium dodecyl sulfate‐polyacrylamide gel electrophoresis. Polynomial regression analysis was used to fit curves to the data, points obtained by plotting log10 of protein molecular weight versus electrophoretic mobility. Polynomials with orders ranging from 1 to 4 were generated. The coefficients of each equation were analyzed for statistical significance. It was found that a third order polynomial was the hightest‐order equation in which all coefficients contributed significantly to the prediction of molecular weights. Using this equation, it was possible to estimate the molecular weights of known proteins in the range from 97 400 to 14 400 with a maximum error of 1 %, compared with a maximum error of 17 % when a first‐order equation was used to describe the migration of the standarads.
Interference of the carbohydrate moiety in Coomassie Brilliant Blue R‐250 protein staining
Authors: Osset Miquel , Piñol Montserrat , Fallon Martin J. M. , De Llorens Rafael , Cuchillo Claudi M. 
Published: May 2005

AbstractThe binding of Coomassie Brilliant Blue R‐250 to several species of bovine pancreatic ribonuclease is affected by the presence of a carbohydrate moiety in the enzyme molecule. Enzymic deglycosylation of several chromatographic fractions of ribonuclease, which have different degrees of glycosylation, results in increased staining by Coomassie Brilliant Blue R‐250. Ovalbumin and other glycoproteins tested show similar behavior. The results indicate that carbohydrate moieties may represent a common hindrance to the binding of Coomassie Brilliant Blue dyes to glycoproteins.
Pulsed field gel electrophoresis: Studies of DNA migration made with the programmable, autonomously‐controlled electrode electrophoresis system
Authors: Birren Bruce W. , Hood Leroy , Lai Eric 
Published: May 2005

AbstractWe have studied the migration of DNA in pulsed field agarose gels under a variety of electrophoresis conditions. We have made use of an instrument which can generate electric fields of any orientation, magnitude, or duration to compare different separation techniques for DNA molecules of from 1 to several thousand kilobase pairs. We discuss the capabilities of the system and present results of gel runs in which electrophoresis conditions were changed individually or in combination. The mobility of DNA in pulsed field gels is shown to reflect a number of interdependent physical parameters.
Separation of chromosomal length DNA molecules: Pneumatic apparatus for rotating gels during electrophoresis
Authors: Sutherland John C. , Emrick Ann B. , Trunk John 
Published: May 2005

AbstractThe maximum length of DNA molecules that can be separated by gel electrophoresis can be increased greatly by periodically altering the direction of the electric field with respect to the gel by an angle that exceeds 90°. One method involves rotating the gel by the desired angle in alternate directions periodically during electrophoresis. We describe a modification of the rotating gel electrophoresis apparatus developed by Serwer (Electrophoresis 1987, 8, 301–304) that uses a pneumatic rotary actuator instead of a stepping motor, hence reducing the cost by about 50 %. Other advantages of our design are a lower center of gravity that makes the apparatus more stable and the removal of all electrical power from beneath the fluid‐filled electrophoresis chamber. We present data demonstrating the separation of chromosomal length DNA molecules from Saccharomyces cerevisiae strain 334 into 14 resolved bands in parallel lanes.
Gel electrophoretic analysis of DNA branched junctions
Authors: Seeman Nadrian C. , Chen Jung‐Huei , Kallenbach Neville R. 
Published: May 2005

AbstractGel electrophoresis has provided much of the detailed information we have about the properties of DNA junctions, stable branched molecules formed from oligonucleotide or polynucleotide strands. Here we review these applications, and present the results of an electrophoretic investigation of conformationally restricted junctions formed by covalently connecting two different pairs of strands in a junction with four arms. Native gel electrophoresis is employed to establish the formation and stoichiometry of the multistrand complexes. Ferguson analysis of native gel mobility shows that junctions have retardation coefficients that are distinct from those of linear DNA duplexes.Denaturing gel electrophoresis is the primary tool for characterizing junctions that have been covalently linked together to form both linear and macrocyclic oligomers of junctions (oligojunctions). Radioactively labelled strands enable one to monitor the progress of the ligation reaction: both linear and closed cyclic molecules result, and these can be distinguished by applying Ferguson analysis to denaturing gels. Combinations of exonuclease III, restriction enzymes and sequencing reactions have been applied to oligojunction molecules, and the results are all analyzed on denaturing gels.Junctions containing intramolecular “tethers” that restrict the conformation freedom of the complex comprise a new system for analyzing the conformations of branched molecules. In these tethered junctions, the ability of arms to moverelative to each other is restricted substantially by covalently connecting pairs of arms in the original complex with short, flexible loops. The two tethers used here constrain the helical domains of the structure to be roughly parallel or anti‐parallel. In this article we use Ferguson analysis to compare two tethered junctions with an untethered junction. At high gel concentrations, the mobility of the untethered complex is found to be closer to that of the molecule tethered anti‐parallel than to the one tethered parallel. Curvature in the Ferguson plots for all three of these junctions is detected over arange of compositions. At low gel concentrations, differences in electrophoretic mobility persist, suggesting that the untethered junction differs in charge as well as conformational freedom from the tethered analogs. We expect that studies of this kind will able to define the conformational repertoire of junctions of different kinds, and to explore the effects of electrophoresis on these states.
Studies of DNA‐protein interactions by gel electrophoresis
Authors: Ceglarek John A. , Revzin Arnold 
Published: May 2005

AbstractThe use of gel electrophoresis in studies of nucleic acid‐protein (especially DNA‐protein) interactions has yielded much qualitative and quantitative information about a variety of such systems. The reduction in mobility of complexes relative to free DNA allows isolation and characterization of the complexes as well as determination of thermodynamic and kinetic properties of the interactions. This article begins with a review of recent applications of the “gel retardation” assay, by way of introduction to experiments in two areas. In the first, a hypothesis is tested regarding whether a DNA molecule with sizable proteins bound very near to each end migrates through a polyacrylamide gel differently than does the corresponding complex having the proteins in the middle of the DNA fragment. The data show little mobility differences for these types of complexes, implying that both may move in a linear, “snakelike”, manner through the gel. The experiments also provide results pertaining to questions of DNA bending caused by the binding of the E. coli catabolite activator protein (CAP) and RNA polymerase to the lactose promoter region. It appears that DNA bending by CAP at its wild type lac binding site is retained in complexes where RNA polymerase is bound simultaneously at the lac UV5 promoter.
The use of denaturing gradient gel electrophoresis to screen for DNA sequence polymorphisms in the human factor VIII gene
Authors: Collins Mary , Wolf Stanley F. , Haines Lora L. , Mitsock Lisa 
Published: May 2005

AbstractWe describe the use of denaturing gradient gel electrophoresis to screen for DNA sequence polymorphisms in the human factor VIII gene. DNA fragments that differ in sequence by only a single base pair can be separated on denaturing gradient gels due to changes in their melting behavior. Previous studies have demonstrated the use of denaturing gradient gels to detect sequence changes in human genomic DNA, including mutations in the β globin gene and polymorphisms on chromosome 20. We have begun to use denaturing gradient gels to look for polymorphisms within the human factor VIII gene. The DNA sequences of seven cloned fragments from introns in the human factor VIII gene were determined and used to predict a melting map for each fragment. The melting behavior of each cloned fragment was evaluated by electrophoresis into denaturing gradient gels. Appropriate fragments were then used as radioactive probes for hybridization to human DNA samples that had been digested with restriction enzymes. Heteroduplexes formed between the probe and genomic DNA samples were electrophoresed into denaturing gradient gels. The final positions of heteroduplex bands were determined by autoradiography. We describe a general approach for using denaturing gradient gel electrophoresis to find DNA polymorphisms, with particular emphasis on the predictive value of DNA sequence data. We compare the efficiency of polymorphism detection by denaturing gradient gel electrophoresis with detection by restriction fragment length polymorphism (RFLP) analysis. The factor VIII gene appears to have a low level of DNA sequence polymorphism. We detected only two rare sequence variants using seven different probes to screen almost 2500 base pairs from each of at least 12 different chromosomes. This low level of sequence polymorphism is consistent with other measurements suggesting that nucleotide variations occur less frequently on the X chromosome than on autosomes.
Reptation‐Breathing theory of pulsed electrophoresis: Dynamic regimes, antiresonance and symmetry breakdown effects
Authors: Viovy Jean L. 
Published: May 2005

AbstractWe apply the concepts of tube and reptation to the pulsed electrophoresis of DNA, considering both biased reptation and “breathing” modes (internal modes of the chain). Using suitable preaveraging approximations, analytical expressions are derived which relate displacement in crossed field electrophoresis to molecular weight, field strength, field period, pore size of the gel, and the angle between the field. These expressions provide scaling laws for the change of mobility when one (or more) of the parameters is varied as well as “universal” velocity versus molecular weight versus pulse time curves. These results are quantitatively compared with experiments. At some point which depends on field angle, field strength and chain length, however, we predict a failure of this model due to symmetry breakdown and loss of ergodicity. Qualitatively, this should lead to considerable band spreading and/or splitting of the highest DNA bands into two bands migrating sideways from the diagonal. The case of field inversion is also investigated. It is shown that only breathing modes can explain the strong differences in mobility experienced by chains of different length when opposite fields of equal amplitude are applied: the “trapping” of chains in conformations of low mobility is associated with an antiresonance‐like coupling between the external field and the internal modes.
Elevated plasma levels of haptoglobin in duchenne muscular dystrophy: Electrophoretic variants in patients with a severe form of the disease
Authors: John Huw A. , Purdom Ian F. 
Published: May 2005

AbstractAnalysis of plasma proteins of Duchenne, muscular dystrophy (DMD) patients and normal age‐matched boys using two dimensional gel electrophoresis and densitometry indicated a three‐fold increase in the average haptoglobin level in DMD plasma. Two electrophoretic variants of haptoglobin in which the α2 chain had more basic or more acidic spots were found in DMD patients with a rapidly progressing form of the disease. The basic variant was present in relatively low amount. Possible reasons for the elevated haptoglobin levels are discussed.
Transparency of nitrocellulose membranes with triton X–114
Authors: Vachereau Andre 
Published: May 2005

AbstractTransparency of nitrocellulose membranes can be realized by simply impregnating the filters with concentrated Triton X‐114. Within a few seconds, protein blots become transparent and can be photographed by transillumination or scanned on a densitometer. Removal of the detergent is possible and the recovered filters can further be used for immunodetection purposes. This procedure is particularly well‐suited where extensive investigation of a single separated protein sample is required.
A two‐dimensional electrophoresis procedure for single meristems of different forest species
Authors: Bon Marie‐Claude 
Published: May 2005

AbstractHigh resolution two‐dimensional electrophoresis, with isoelectric focusing in the first dimension and electrophoresis in sodium dodecyl sulfate in thin acrylamide gels in the second dimension, has been applied to separate the proteins of single meristems (200‐300 μm) from Sequoiadendrongiganteum, Sequoiasempervirens, Pseudotsuga menziesii, Picea abies, Pinus pinaster, Eucalyptus gunnii and Populus nigra. The technique may prove an efficient tool for studying inter‐ and intraclonal differences in meristem cultures of forest species, especially when referring to cloning from meristems of selected trees in relation to the rejuvenation process.
Two‐dimensional electrophoretic analysis of nuclear proteins from human tumors
Authors: Menzel Alain , Unteregger Gerhard 
Published: May 2005

AbstractDuring the last decade several strategies have been developed to identify proteins which could serve as markers in tumor biology. One avenue of great promise to detect such proteins seems to be the separation of prefractionated organelles from tumor cells by high resolution two‐dimensional electrophoresis. Using detergent‐lysed nuclei from several human tumor cell lines, especially from brain tumors, and two‐dimensional electrophoresis, we analyzed the nuclear protein pattern obtained after sequential salt extraction of tumor cell nuclei. In addition to proteins occurring in all tumor cell lines, the pattern of different tumor cell lines exhibits considerable differences when proteins were visualized by silver staining, thus emphasizing the specificity of nuclear proteins with respect to the cell type. Even quantitative variations of the nuclear phosphoproteins 23/4 were detectable, indicating a potential correlation between their synthesis/phosphorylation and the proliferation behavior of tumor cells. The data indicate that nuclear proteins with their distinct heterogeneity and tissue specificity may represent a powerful source in determining tumor‐specific proteins. The extent of chromosomal protein heterogeneity may be additionally increased by their covalent modification by nuclear kinases; therefore, tumor‐specific nuclear proteins may occur as quantitative and qualitative variations.
Lectin affinity electrophoresis of α‐fetoprotein in cancer diagnosis
Authors: Taketa Kazuhisa , Hirai Hidematsu 
Published: May 2005

AbstractLectin affinity electrophoresis of serum α‐fetoprotein (AFP) was carried out on samples obtained from patients with benign and malignant diseases and on cord blood, and separated AFP bands were detected by antibody‐affinity blotting. The following major bands were identified by determination of kinetic constants: AFP‐C1 and ‐C2 with concanavalin A, AFP‐LI, ‐L2 and ‐L3 with Lens culinaris agglutinin A, AFP‐P1, ‐P2, ‐P3, ‐P4 and ‐P5 with erythroagglutinating phytohemagglutinin, and AFP‐A1, ‐A2 and ‐A3 with Allomyrina dichotoma lectin. AFP bands with the lowest number had either low or no affinity and those with higher numbers had higher affinities for respective lectins. AFP from cord blood and chronic liver disease was characterized by the predominance of AFP‐C2, AFP‐L1, AFP‐P2 and AFP‐A3. Hepatocellular carcinoma was differentiated from the benign liver disease by increased proportions of AFP‐L3 and AFP‐P4. Extrahepatic tumors had additional increases of AFP‐C1, AFP‐L2, AFP‐P5 and AFP‐A1 (or slow‐migrating AFP‐Als, particularly in yolk sac tumor).
Electrophoretically silent hemoglobin mutants as revealed by isoelectric focusing in immobilized pH gradients
Authors: Righetti Pier Giorgio , Gianazza Elisabetta , Bianchi‐Bosisio Adriana , Wajcman Henri , Cossu Gianfranco 
Published: May 2005

AbstractThe applications of isoelectric focusing in immobilized pH gradients to the analysis of (i) human hemoglobin mutants, (ii) animal hemoglobin mutants (from cattle, sheep, dog and mouse), and (iii) tryptic digests of α and β chains, are discussed and evaluated. Immobilized pH gradients appear to be an excellent tool for screening of genetic polymorphism and for detecting “silent mutants”,i. e. those substitutions involving amino acids with nonionizable side chains. At present, not even capillary zone electrophoresis, claimed to have a resolving power equivalent to 1 million theoretical plates, has shown a resolution capability comparable to that of immobilized pH gradients, at least in the field of protein separation.