bluuebunny/crossref_metadata_2025 huggingface dataset:
Perspectives and limitations of resolutions–reconstitution experiments
Authors: Racker Efraim
Published: May 2005

Reconstitutions of membranous activities can tell us how many components are required and what their functions are. The mitochondrial proton pump is used as an example. Moreover, the biological activity, such as Pi transport, can be used in reconstituted vesicles as an assay during the isolation of the transporter.

Reconstitution experiments reveal the importance of membrane asymmetry and allow us to study conditions of vectorial assembly.

The mechanism of action of ion pumps has been successfully analyzed in reconstituted liposomes. We can study the movement of ions and the electrogenicity of the system without interference by other unrelated processes.

Based on studies with the resolved Ca2+‐ATPase of sarcoplasmic reticulum, we propose a novel formulation of the mechanism of ATP‐driven ion pumps in which cyclic binding of Mg2+ plays a key role.
Plants interact with microbial polysaccharides
Authors: Albersheim Peter , Ayers Arthur R. , Valent Barbara S. , Ebel Jürgen , Hahn Michael , Wolpert Jack , Carlson Russell
Published: May 2005

Plants are resistant to almost all of the microorganisms with which they come in contact. In response to invasion by a fungus, bacterium, or a virus, many plants produce low molecular weight compounds, phytoalexins, which inhibit the growth of microorganisms. Phytoalexins are produced whether or not the invading microorganism is a pathogen. The production of phytoalexins appears to be a widespread mechanism by which plants attempt to defend themselves against pests. Molecules of microbial origin which trigger phytoalexin accumulation in plants are called elicitors. Structural polysaccharides from the mycelial walls of several fungi elicit phytoalexin accumlation in plants. Approximately 10 ng of the polysaccharide elicits the accumulation in plants of more than sufficient amounts of phytoalexin to stop the growth of microorganisms in vitro. The best characterized elicitors have been demonstrated to be β‐1,3‐glucans with branches to the 6 position of some of the glucosyl residues. Oligosaccharides, produced by partial acid hydrolysis of the mycelial wall glucans, are exceptionally active elicitors. The smallest oligosaccharide which is still an effective elicitor is composed of about 8 sugar residues.

Bacteria also elicit phytoalexin accumulation in plants, but the Rhizobium symbionts of legumes presumably have a mechanism which allows them to avoid either eliciting phytoalexin accumulation or the effects of the phytoalexins if they are accumulated. The lectins of legumes bind to the lipopolysaccharides of their symbiont, but not of their non‐symbiont, Rhizobium. It is not known whether the lectin‐lipopolysaccharide interaction is involved with the establishment of symbiosis. However, evidence will be presented that suggests that lectins are, in fact, enzymes capable of modifying the structurs of the lipopolysaccharides of their symbiont, but not of their non‐symbiont, Rhizobium. It will also be shown that the lipopolysaccharides isolated from different Rhizobium species and from different strains of individual Rhizobium species have different sugar compositions. Thus, the different strains of a single Rhizobium species are as different from one another as the different species of Salmonella and other gram‐negative bacteria. This conclusion is substantiated by experiments demonstrating that antibodies to the lipopolysaccharide from a single Rhizobium strain can differentiate that strain from other strains of the same species as well as from other Rhizobium species. The role in symbiosis of the strain‐specific O‐antigens is unknown.
Studies related to the head‐maturation pathway of bacteriophages T4 and T2: I. Morphology and kinetics of intracellular particles produced by mutants in the maturation genes
Authors: Wunderli H. , v.d. Broek J. , Kellenberger E.
Published: May 2005

Mutants in the genes governing the maturation of the head of bacteriophage T4 and in gene 24 were studied by electron microscopy of thin sections. We define morphologically: black particles, comprising mature, stable heads and immature, fragile heads, which break down upon lysis; grizzled particles, which apparently are partially filled or partially emptied; empty large particles without DNA or core Which are all the same size as normal heads; empty small particles without DNA and without core which are of the size of the τ particle, which is the prehead of phage T4.

The study of single and double mutants of the maturation genes demonstrates that the phenotypes are only different by the proportions of the different particles made except for 17− where only empty small and empty large particles accumulate. The mutants in gene 24 are epistatic on all other mutants. Mutants in gene 17 are epistatic on the remaining ones.

The results are consistent with the hypothesis that the products of several of the maturation genes act on DNA to render it competent for packaging while the others act directly on the particle. By this uncoupling, bypasses and abortive pathways can result.
Effect of calcium on the pellet height response of Tetrahymena cilia
Authors: Blum J. J. , Hayes Alvernon
Published: May 2005

The pellet height response (a measure of the increase in height of the pellet of cilia obtained by brief centrifugation in the presence of ATP as compared to the absence of ATP) of Tetrahymena cilia prepared by deciliation in the presence of Ca2+ is sensitive to the concentration of free Ca2+ during the pellet height assay. The magnitude of the increase in pellet height and the sharpness of the pellet boundary both increase markedly with increasing [Ca2+]. The half‐maximal effect is attained at a free [Ca2+] of about 1.5 × 10−7 M. The pellet height assay thus measures a Ca2+‐sensitive component of the ciliary motile system. The possibility that this is the Ca2+‐sensitive orientation system is discussed.
Reconstitution of D‐glucose transport in vesicles composed of lipids and intrinsic protein (Zone 4.5) of the human erythrocyte membrane
Authors: Kahlenberg Arthur , Zala Cedric A.
Published: May 2005

Elucidation of the mechanism of facilitated D‐glucose transport in human erythrocytes is dependent on the identification and isolation of the membrane protein(s) mediating this process. Based on the fact that stereospecific D‐glucose transport is reconstituted in liposomes prepared by sonication of a lipid suspension with ghosts or fractions derived from ghosts, a quantitative assay for the stereospecific D‐glucose transport activity of these fractions was developed (Zala CA, Kahlenberg A: Biochem Biophys Res Commun 72:866, 1976). This assay was used to monitor the purification of ghosts. The solubilized membrane protein fraction was chromatographed on a column of diethylaminoethyl cellulose which was eluted stepwise with NaCl‐phosphate buffers of increasing ionic strength. A fraction, eluted at an ionic strength of 0.1, displayed a 13‐ and 27‐fold increase in reconstituted transport activity relative to ghosts and to the unfractionated Triton X‐100 extract, respectively. This fraction, when analyzed by sodium dodecyl sulfate‐polyacrylamide gel electrophoresis, consisted predominantly of the ghost proteins with an apparent molecular weight of 55,000, commonly designated as zone 4.5; periodic acid‐Schiff‐sensitive membrane glycoproteins 1–4 were absent. Transport reconstituted by this preparation of zone 4.5 membrane proteins was almost completely abolished by 1‐fluoro‐2,4‐dinitrobenzene, mercuric chloride, and p‐chloromercuribenzene sulfonate, but was unaffected by sodium iodoacetate. Extra‐ and intraliposomal phloretin and cytochalasin B, respectively, exhibited partial inhibition. The stereospecificity and inhibition characteristics of the reconstituted transport imply that all the components of the erythrocyte D‐glucose transport system are contained in the zone 4.5 membrane protein preparation.
The molecular mechanism of dicarboxylic acid transport in escherichia coli K 12
Authors: Lo Theodore C. Y.
Published: May 2005

It is the purpose of this communication to review the properties of the dicarboxylic acid transport system in Escherichia coli K12, in particular the role of various dicarboxylate transport proteins, and the disposition of these components in the cytoplasmic membrane. The dicarboxylate transport system is an active process and is responsible for the uptake of succinate, fumarate, and malate. Membrane vesicles prepared from the EDTA, lysozyme, and osmotic shock treatment take up the dicarboxylic acids in the presence of an electron donor. Genetic analysis of various transport mutants indicates that there is only one dicarboxylic acid transport system present in Escherichia coli K12, and that at least 3 genes, designated cbt, dct A, and dct B, are involved in this transport system. The products corresponding to the 3 genes are: a periplasmic binding protein (PBP) specified by cbt, and 2 membrane integral proteins, SBP 1 and SBP 2, specified by dct B and dct A, respectively. Components SBP 1 and SBP 2 appear to be exposed on both the inner and outer surfaces of the membrane, and lie in close proximity to each other. The substrate recognition sites of SBP 2 and SBP 1 are exposed on the outer and inner surfaces of the membrane respectively. The data presently available suggest that dicarboxylic acids may be translocated across the membrane via a transport channel. A tentative working model on the mechanism of translocation of dicarboxylic acids across the cell envelope by the periplasmic binding protein, and the 2 membrane carrier proteins is presented.
Existence of a common precursor to ACTH and endorphin in the anterior and intermediate lobes of the rat pituaitary
Authors: Eipper Betty A. , Mains Richard E.
Published: May 2005

Extracts of rat anterior and intermediate‐posterior pituitary were fractionated by sodium dodecyl sulfate polyacrylamide gel electrophoresis and assayed for immunoactive ACTH and endorphin. In both lobes the major forms of immunoactive ACTH have apparent molecular weights of 31,000 (31K), 20–21K, 14K, and 4.5K, and the major forms of immunoactive endorphin have apparent molecular weights of 31K (coincident with the peak of immunoactive ACTH), 13K (a βLPH‐like peptide), and 3.5K (a β‐endorphin‐like peptide). However, the quantitative distribution of immunoactivity among the various forms differs greatly between the lobes. Assays using an extreme COOH‐terminal ACTH antiserum indicate that the 31K ACTH/endorphin molecule in rat antierior and intermediate pituitary is similar to the pro‐ACTH/endorphin molecule from mouse pituitary tumor cells. A radioimmunoassay that is specific for the NH2‐terminal non‐ACTH, nonendorphin segment (referred to as 16K fragment) of the mouse pro‐ACTH/endorphin molecule was used to assay extracts of rat pituitary. In addition to detecting material at 31K and 20–21K, the 16K fragment radioimmunoassay detects significant amounts of cross‐reactive material with an apparent molecular weight of 16K in extracts of both lobes. This result also suggests that the structure and processing of the rat 31K ACTH/endorphin molecule is similar to that of mouse tumor cell pro‐ACTH/endorphin.

Cell suspensions were prepared from the anterior and intermediate lobes of the rat pituitary and maintained in culture for a 24‐h period. The isolated cells from both lobes incorporate [3H] phenylalanine into immunoprecipitable ACTH‐ and endorphin‐containing molecules. By sequential immunoprecipitation with ACTH and endorphin antisera, it is possible to demonstrate directly that a single molecule (31K ACTH/endorphin) has antigenic determinants for both ACTH and endorphin. Significant amounts of 31K ACTH/endorphin are released into the culture medium by isolated anterior lobe and intermediate lobe cells. The isolated intermediate lobe cells synthesize and secrete relatively large amounts of a β‐endorphin‐like molecule; the isolated anterior lobe cells secrete significant amounts of both a βLPH‐like molecule and a β‐endorphin like molecule. These same quantitative differences between anterior and intermediate lobe tissue were observed in immunoassays of extracts of the separated lobes and probably reflect differences in the processing of the common precursor. The isolated anterior lobe cells can be stimulated to release increased amounts of immunoprecipitable ACTH and endorphin by incubation with a cyclic AMP analog and a phosphodiesterase inhibitor.
Fine structure of the band 3 protein in human red cell membranes: Freeze‐fracture studies
Authors: Weinstein Ronald S. , Khodadad Jena K. , Steck Theodore L.
Published: May 2005

The major red cell membrane protein, band 3, is a glycoprotein which extends across the membrane from the extracellular space into the cytoplasmic compartment. It is widely held that band 3 is a component of the intramembrane particles (IMP) which can be demonstrated by freeze‐fracture electron microscopy. In this study, we find that the outer surface poles of the IMP can be seen by freeze‐etching after they are unmasked by proteolysis under conditions which excise the surrounding sialopeptides from the membrane. The poles appear as distinctive projections, 30–50 Å in diameter, the “ES particles.” The ES particles remain associated with the outer surface of the membrane following cleavage of the band 3 polypeptide by chymotrypsin or pronase. This is consistent with previous biochemical studies which have shown that the 38,000‐dalton outer surface segment of band 3 is intercalated in the lipid bilayer. A granulofibrillar component at the inner surface of the membrane is provisonally identified as the 40,000‐dalton inner‐surface domain of band 3.
Glucocorticoid‐mediated alteration in growth factor binding and action: Analysis of the binding change
Authors: Baker Joffre B. , Cunningham Dennis D.
Published: May 2005

The addition of the glucocorticoid analog dexamethasone (DX) to serum‐free cultures of human fibroblasts caused a twofold enhancement of the mitogenic response to epidermal growth factor (EGF), although DX by itself was not mitogenic. A basis for this effect was suggested by studies showing that DX also increased the cellular binding of 125I‐EGF. DX increased the ability of the cells to bind 125I‐EGF only at low physiological concentrations of this polypeptide. Thus, data from 125I‐EGF binding to cells incubated without DX produced a linear Scatchard plot, whereas the data from 125I‐EGF binding to DX‐treated cells led to an upwardly curvilinear Scatchard plot. Measurements of 125I‐EGF association with the cell surface and cytoplasm indicated that this binding change involved an alteration of cell surface EGF receptors. The binding change appeared not to involve negatively cooperative interactions between EGF receptors, nor a change in the number of receptors. The binding alteration could be explained by a model in which DX converted 25–30% of the cell surface EGF receptors to a form having a fourfold increased affinity.
Proteolytic cleavage and structural transformation: Their relationship in bacteriophage T4 capsid maturation
Authors: Steven Alasdair C. , Carrascosa Jose L.
Published: May 2005

Giant T4 phage capsoids formed in canavanine‐treated cultures infected by phage mutants in genes 21 and 17, respectively, differ with regard to cleavage of the major capsid protein, gp 23, and in the fine structure of their hexagonal surface lattices. Quantitative computer processing of electron micrographs shows that the significant differences in capsomer morphology amount to six symmetrically placed features present in the uncleaved hexamer but absent after cleavage. These features may be related with the N‐terminal portions of gp 23 monomers excised by phage‐specific proteolysis. Cleaved 17− giants can be induced to undergo a further structural transformation (expansion). Structural characteristics of partially transformed giant particles give clues about the dynamics of the cleavage and expansion transformations. Both processes appear to be polar, initiating in one cap and propagating along the particle. The transition zone of partial cleavage is diffuse, whereas the transition between unexpanded and expanded areas is confined to a narrow band of some 20 nm width.
Growth factors and gangliosides: A possible new perspective in neuronal growth control
Authors: Morgan James I. , Seifert Wilfried
Published: May 2005

For many permanent cell lines the transition from a growing (P) to a resting (R) state is reversibly controlled by growth factors present in serum. This P‐to‐R transition was studied in a neuronal cell line (B 104) with respect to the action of serum, dibutyryl cyclic AMP (DBcAMP), gangliosides, and a glioma cell‐produced growth factor GGF. In this cell system gangliosides seem to act as differentiation and survival factors. The kinetics of uptake of radioactively labeled gangliosides and survival experiments both support the idea of the stable incorporation of exogenously added gangliosides into the cells. Based on the experimental evidence a new model of cell development is proposed. Thus in addition to the R or G0 state, which in this cell system is rather unstable and probably regulated by cyclic nucleotides, we postulate a differentiated D state, which is controlled by gangliosides and which is characterized by its stability (survival time). This D compartment seems to be closer to the in vivo differentiated neuron than does the R or P state. The possible mechanisms for the action of gangliosides are discussed.
Co‐translational modification of nascent immunoglobulin heavy and light chains
Authors: Bergman Lawrence W. , Kuehl W. Michael
Published: May 2005

We have investigated the in vivo co‐translational covalent modification of nascent immunoglobulin heavy and light chains. Nascent polypeptides were separated from completed polypeptides by ion‐exchange chromatography of solubilized ribosomes on QAE‐Sephadex. First, we have demonstrated that MPC 11 nascent heavy chains are quantitatively glycosylated very soon after the asparaginyl acceptor site passes through the membrane into the cisterna of the rough endoplasmic reticulum. Nonglycosylated completed heavy chains of various classes cannot be glycosylated after release from the ribosome, due either to rapid intramolecular folding and/or intermolecular assembly, which cause the acceptor site to become unavailable for the glycosylation enzyme. Second, we have shown that the formation of the correct intrachain disulfide loop within the first light chain domain occurs rapidly and quantitatively as soon as the appropriate cysteine residues of the nascent light chain pass through the membrane into the cisterna of the endoplasmic reticulum. The intrachain disulfide loop in the second or constant region domain of the light chain is not formed on nascent chains, because one of the cysteine residues involved in this disulfide bond does not pass through the endoplasmic reticulum membrane prior to chain completion and release from the ribosome. Third, we have demonstrated that some of the initial covalent assembly (formation of interchain disulfide bonds) occurs on nascent heavy chains prior to their release from the ribosome. The results are consistent with the pathway of covalent assembly of the cell line, in that completed light chains are assembled onto nascent heavy chains in MPC 11 cells (IgG2b), where a heavy‐light half molecule is the major initial covalent intermediate; and completed heavy chains are assembled onto nascent heavy chains in MOPC 21 cells (IgG1), where a heavy chain dimer is the major initial disulfide linked intermediate.
The detection, immunofluorescent localization, and thrombin induced release of human platelet‐associated fibronectin antigen
Authors: Ginsberg Mark H. , Painter Richard G. , Birdwell Charles , Plow Edward F. , Forsyth Jane
Published: May 2005

Platelets are cells which develop adhesive properties following stimulation. Since fibronectin (fn) mediates adhesive properties of several cells, we sought evidence for platelet associated fn. Lysates of suspensions of washed human platelets containing ≤50 ng soluble fn/109 cells contained 2.85 μg fn antigen per 109 cells. The platelet fn antigen competition curve showed a similar slope to the curve for purified plasma fn suggesting antigenic identity. Immunofluorescent staining for fn was minimal in intact cells suggesting that the majority of fn antigen is intracellular. In permeable platelets, fluorescent staining for fn was seen in a punctate distribution suggesting a granule localization. Stimulation of platelet secretion by thrombin released platelet fn antigen. Suramin, a drug which inhibits platelet secretion, inhibited fn release. The apparent secretion of platelet fn, taken with the immunofluorescent data, support the localization of a portion of platelet fn antigen in a storage granule.
Fibronectin and proteoglycans as determinants of cell‐substratum adhesion
Authors: Culp Lloyd A. , Murray Ben A. , Rollins Barrett J.
Published: May 2005

When normal or SV40‐transformed Balb/c 3T3 cells are treated with the Ca++‐specific chelator EGTA, they round up and pull away from their footpad adhesion sites to the serum‐coated tissue culture substrate, as shown by scanning electron microscope studies. Elastic membranous retraction fibers break upon culture agitation, leaving adhesion sites as substrate‐attached material (SAM) (Cells leave “footprints” of substrate adhesion sites during movement by a very similar process.) SAM contains 1–2% of the cell's total protein and phospholipid content and 5–10% of its glucosamine‐radiolabeled polysaccharide, most of which is glycosaminoglycan (GAG). By one‐ and two‐dimensional sodium dodecyl sulfate‐polyacrylamide gel electrophoresis, there is considerable enrichment in SAM for specific GAGs; for the glycoprotein fibronectin; and for the cytoskeletal proteins actin, myosin, and the subunit protein of the 10 nm‐diameter filaments. Fibrillar fibronectin of cellular origin and substratum bound fibronectin of serum origin (cold‐insoluble globulin, CIg) have been visualized by immunofluorescence microscopy. The GAG composition in SAM has been examined under different cellular growth and attachment conditions. Heparan sulfate content correlates with glycopeptide content (derived from glycoprotein). Newly attaching cells deposit SAM with principally heparan sulfate and fibronectin and little of the other GAGs. Hyaluronate and chrondroitin proteoglycans are coordinately deposited in SAM as cells begin spreading and movement over the substrate. Cells attaching to serum‐coated or CIg coated substrates deposited SAM with identical compositions. The proteoglycan nature of the GAGs in SAM has been examined as well as the ability of proteoglycans to form two classes of reversibly dissociable “supramolecular complexes” ‐ one class with heparan sulfate and glycopeptide‐containing material and the second with hyaluronate‐chondroitin complexes. Enzymatic digestion of “intact” SAM with trypsin or testicular hyaluronidase indicates that (1) only a small portion of long‐term radiolabeled fibronectin and cytoskeletal protein is bound to the substrate via hyaluronate or chondroitin classes of GAG; (2) most of the fibronectin, cytoskeletal protein and heparan sulfate coordinately resist solubilization; and (3) newly synthesized fibronectin, which is metabolically labile in SAM, is linked to SAM by hyaluronate‐ and/or chondroitin‐dependent binding. All of our studies indicate that heparan sulfate is a direct mediator of adhesion of cells to the substrate, possibly by binding to both cell‐surface fibronectin and substrate‐bound CIg in the serum coating; hyaluronate‐chondroitin complexes in SAM appear to be most important in motility of cells by binding and labilizing fibronectin at the periphery of footpad adhesions, with subsequent cytoskeletal disorganization.
Possible role of fibronectin in malignancy
Authors: Chen Lan Bo , Summerhayes Ian , Hsieh Philip , Gallimore Philip H.
Published: May 2005

Frozen sections of tumors induced by injecting virally transformed cells into animals were stained for fibronectin by immunofluorescence. Many tumor cell lines do not express fibronectin in tumors in situ even though some of them express fibronection in culture. Cell shape and hormones appear to influence the expression of fibronectin in culture; however, it is nuclear how fibronection expression is regulated in vivo.
Regulation of the Balb/c‐3T3 cell cycle‐effects of growth factors
Authors: Stiles C. D. , Pledger W. J. , Tucker R. W. , Martin R. G. , Scher C. D.
Published: May 2005

The platelet‐derived growth factor (PDGF), which is found in serum but not in plasma, has been purified to homogeneity; it stimulates replication at a concentration of 10−10M. Brief treatment with PDGF causes densityinhibited Balb/c‐3T3 cells to become competent to synthesize DNA; pituitary fibroblast growth factor (FGF) or precipitates of calcium phosphate also induce competence. Continuous treatment with plasma allows competent, but not incompetent, cells to synthesize DNA. A critical component of plasma is somatomedin, a group of hormones with insulin‐like activity; multiplication‐stimulating activity (MSA) or insulin replace plasma somatomedin in promoting DNA synthesis.

We have studied the molecular correlates of competence and the role of SV40 gene A products in regulating DNA synthesis. Treatment of quiescent cells with pure PDGF or FGF causes the preferential synthesis of five cytoplasmic proteins (approximate molecular weight 29,000, 35,000, 45,000, 60,000, and 72,000 detected by SDS‐PAGE under reducing conditions). Two of these competence‐associated proteins (29,000 and 35,000 daltons) are found within 40 min of PDGF addition; they are not induced by plasma, insulin, or epidermal growth factor (EGF), PDGF, FGF, or calcium phosphate induce an ultrastructure change within the centriole of 3T3 cells; this ultrastructural modification of the centriole is detectable by immunofluorescence within 2 h of PDGF treatment. Plasma, EGF, or MSA do not modify the centriole. SV40 induces replicative DNA synthesis in growth‐arrested 3T3 cells but does not cause this alteration in centriole structure.

Gene A variants of SV40, including a mutant with temperature‐sensitive (ts) T‐antigen (ts A209), a deletion in t‐antigen (dl 884), and several ts A209 strains containing t‐antigen deletions were used to induce DNA synthesis in Balb/c‐3T3 cells. Like wild type SV40, all strains induced DNA synthesis equally well under permissive or nonpermissive conditions. Addition of PDGF or plasma had little effect on SV40‐induced DNA synthesis. Thus, the viral function that induces replicative DNA synthesis in Balb/c‐3T3 cells is not t and is not temperature sensitive. This SV40 gene function overrides the cellular requirement for hormonal growth factors. It does not induce transient centriole deciliation, a hormonally regulated event.
Multiplication stimulating activity (MSA) from the BRL 3A rat liver cell line: Relation to human somatomedins and insulin
Authors: Rechler Matthew M. , Nissley S. Peter , King George L. , Moses Alan C. , Van Obberghen‐Schilling Ellen E. , Romanus Joyce A. , Knight Alfred B. , Short Patricia A. , White Robert M.
Published: May 2005

The properties of multiplication stimulating activity (MSA), an insulin‐like growth factor (somatomedin) purified from culture medium conditioned by the BRL 3A rat liver cell line are summarized. The relationship of MSA to somatomedins purified from human and rat plasma are considered. MSA appears to be the predominant somatomedin in fetal rat serum, but a minor component ot adult rat somatomedin. In vitro biological effects of MSA and insulin in adipocytes, fibroblasts and chondrocytes are examined to determine whether they are mediated by insulin receptors or insulin‐like growth factor receptors. The possible relationship of a primary defect of insulin receptors observed in fibroblasts from a patient with the rare genetic disorder, leprechaunism, to intrauterine growth retardation is discussed.
Monoclonal antibody (M2) to glial and neuronal cell surfaces
Authors: Lagenaur C. , Schachner M.
Published: May 2005

A monoclonal antibody designated M2 arose from the fusion of mouse myeloma cells with splenocytes from a rat immunized with particulate fraction from early postnatal mouse cerebellum. Expression of M2 antigen was examined by indirect immunofluorescence on frozen sections of developing and adult mouse cerebellum and on monolayer cultures of early postnatal mouse cerebellar cells. In adult cerebellum, M2 staining outlines the cell bodies of granule and Purkinje cells. A weaker, more diffuse staining is seen in the molecular layer and white matter. In sections of newborn cerebellum, M2 antigen is weakly detectable surrounding cells of the external granular layer and Purkinje cells. The expression of M2 antigen increases during development in both cell types, reaching adult levels by postnatal day 14. At all stages of postnatal cerebellar development, granule cells that have completed migration to the internal granule layer are more heavily stained by M2 antibodies than are those before and in process of migration. In monolayer cultures, M2 antigen is detected on the cell surface Of all GFA protein‐positive astrocytes and on more immature oligodendrocytes, that express 04 antigen but not 01 antigen. After 3 days in culture, tetanus toxinpositive neurons begin to express M2 antigen. The same delayed expression of M2 antigen on neurons is observed in cultures derived from mice ranging in age from postnatal day 0 to 10.
Presence of trifoliin A, a rhizobium‐binding lectin, in clover root exudate
Authors: Dazzo Frank B. , Hrabak Estelle M.
Published: May 2005

Trifoliin A, a Rhizobium‐binding glycoprotein from white clover, was detected in sterile clover root exudate by a sensitive immunofluorescence assay employing encapsulated cells of Rhizobium trifolii 0403 heat‐fixed to microscope slides. Its presence in root exudate was further examined by immunoaffinity chromatography. The binding of trifoliin A to cells was specifically inhibited by the hapten, 2‐deoxyglucose. Significantly higher quantities of trifoliin A were detected in root exudate of seedlings grown hydroponically in nitrogen‐free medium than in rooting medium containing 15 mM NO, a concentration which completely suppressed root hair infection by the nitrogen‐fixing symbiont. The presence of trifoliin A in root exudate may make it possible for recognition processes to occur before the microsymbiont attaches to its plant host.
Biochemical analysis of an MHC‐linked hematopoietic cell surface antigen, Qa‐2
Authors: Soloski Mark J. , Vitetta Ellen S. , Flaherty Lorraine , Uhr Jonathan W.
Published: May 2005

The region of the murine 17th chromosome telomeric to H‐2D encodes a group of serologically defined cell surface antigens termed Qa‐1‐5. These antigens are of interest because their expression is restricted to hematopoietic cells. In addition, the molecular weight and subunit structure (ie, association with β‐2 microglobulin) of Qa‐2 molecules are similar to H‐2 and TL antigens. In the present studies, we have prepared isotopically labeled Qa‐2 and H‐2 molecules from mitogen‐stimulated C57BL/6 spleen cells. Comparative peptide mapping of tryptic peptides from Qa‐2 and H‐2 molecules (Kb, DbKk, Dd) reveal that Qa‐2 has a unique primary structure. However, considerable homology is indicated since 30–40% of the Qa‐2 peptides cochromatograph with peptides derived from H‐2Kb, H‐2Db, H‐2Kk, and H‐2Dd. Studies by other investigators have demonstrated that similar levels of structural homology are observed when H‐2K, H‐2D, and H‐2L tryptic peptides are analyzed. We conclude from these studies that the Qa‐2 alloantigen is structurally related to a class of cell surface molecules (ie, H‐2) that play critical roles in immune recognition processes. These data further suggest that the genes encoding Qa‐2 and H‐2 molecules have arisen from a common primordial gene.
Monoclonal antibody covalently coupled to liposomes: Specific targeting to cells
Authors: Barbet Jacques , Machy Patrick , Leserman Lee D.
Published: May 2005

We have evaluated optimal conditions for coupling monoclonal antibody to small unilamellar lipisomes. Coupling of an IgG2a monoclonal anti‐β2‐microglobulin antibody, which reacts with human cells, was examined in detail. Liposomes were composed of dipalmitoyl lecithin and cholesterol, and variable quantities of phosphatidylethanolamine substituted with the heterobifunctional cross‐linking reagent N‐hydroxysuccinimidyl 3‐(2‐pyridyldithio) propionate (SPDP). They were reacted with antibody derivatized with the same reagent at a 5‐ to 20‐fold molar excess, and activated by mild reduction. This degree of SPDP modification had no effect on the capacity of the antibody to bind to its target antigen. More than 40% of antibody could be reproducibly bound to liposomes, resulting in the coupling of from 1 to 10 antibody molecules per liposome (mean diameter.580 Å). The coupling reaction did not lead to loss of carboxyfluorescein encapsulated within liposomes. At least 80% of liposomes carried nondenatured antibody, as confirmed by precipitation of liposomes and encapsulated carboxyfluorescein by Staphylococcus aureus, strain Cowan I. The liposome‐coupled antibody retained its immunological specificity: only cells expressing human β2‐microglobulin bound liposomes in vitro, and the binding was inhibited by the free antibody in solution. Results with antibodies of different antigenic specificity confirm that the technique can be generally applied.
Macrophages and methylthio groups in lymphocyte proliferation
Authors: Toohey John I.
Published: May 2005

Macrophages are shown to replace methylthio disulfides in supporting in vitro proliferation of three cell lines previously characterized as methylthio‐dependent. Macrophages have the capacity to generate methylthio groups from methylthioadenosine. It is hypothesized that macrophages stimulate cell proliferation both in normal immune systems and in certain cancers by providing an abundance of methylthio groups. Fetal calf serum is shown to contain methylthio groups. It appears that, in cell cultures containing fetal calf serum, sulfhydryl compounds stimulate cell proliferation by making the methylthio groups in the serum available to the cells.
Fibronectin expression on the platelet surface occurs in concert with secretion
Authors: Ginsberg Mark H. , Plow Edward F. , Forsyth Jane
Published: May 2005

Thrombin stimulation of human platelets causes increased cellular adhesiveness for other platelets (aggregation) and surfaces and increased surface expression of platelet fibronectin antigen. Aggregation occurs concurrently with secretion. In these studies, the threshold thrombin dose for surface expression of fibronectin, as measured by binding of F(ab′)2 antifibronectin, was similar to that for serotonin secretion. Moreover, both processes occurred at similar rates, and inhibition of secretion was associated with inhibition of antifibronectin binding. Thus a hypothesis is proposed in which adhesive proteins within platelet granules become expressed on the platelet surface as a direct consequence of the secretory process. This cluster of adhesive proteins may then contribute to increased cellular adhesiveness.
Spectrin oligomers: A structural feature of the erythrocyte cytoskeleton
Authors: Morrow Jon S. , Haigh Wallace B. , Marchesi Vincent T.
Published: May 2005

Spectrin reversibly self‐associates to high molecular weight oligomers through a concentration‐driven process characterized by association constants of about 105 mol−1. This association is prominent under physiological conditions of pH, ionic strength, and temperature. It is disrupted by urea, but not Triton X‐100. The process of spectrin association appears mathematically to resemble that for tropomyosin, although the mechanism is probably different. Spectrin association is weak compared to other prominent protein–protein associations in the red cell membrane skeleton. The linkage of these weak and strong associations suggests a process whereby the membrane skelton spontaneously assembles. Such affinity‐modulated assembly involving weak associations is likely to be the focus of numerous membrane control mechanisms.
Untersuchungen zur Lochkorrosion (Lochfraß) von hochlegierten austenitischen Chrom‐Nickel‐Stählen und ferritischen Chrom‐Stählen in halogenidhaltigen, wäßrigen Lösungen
Authors: Pier Günther , Schwenk Wilhelm
Published: May 2005

Die Einsatzmöglichkeit der chemisch beständigen Stähle in halogenidhaltigen, wäßrigen Medien wird durch die Erscheinung der Lochfraßkorrosion eingeschränkt.

Es wird gezeigt, daß bei der allgemein üblichen Prüfung in einer 10%igen FeCl3‐Lösung für die Lochfraßanfälligkeit nicht das Korrosionspotential, sondern der Polarisationswiderstand als elektrochemische Meßgröße kennzeichnend ist.

Als praktisches Schnellprüfverfahren hat sich der K3[Fe(CN)6]‐Test bewährt. Mit Hilfe dieser chemischen Indikation wird der Lochfraß‐Mechanismus als Aktiv‐Passiv‐Reaktion bewiesen. Das Auftreten der Lochfraßkorrosion ist an einen gewissen Potentialbereich gebunden, der durch Messung der Stromdichte‐Potential‐Kurve bestimmt werden kann.

In reiner lufthaltiger 5%iger Schwefelsäure ist bei Raumtemperatur nur die gleichmäßig transpassive Stahlauflösung möglich. In chloridhaltiger Schwefelsäure kann der Stahl gleichmäßig aktiv oder durch Lochfraß, dagegen nicht transpassive korrodieren. Durch einen Nitratzusatz zur chloridhaltigen Schwefelsäure wird die Lochfraßkorrosion zugunsten der transpassiven Korrosion inhibitiert.
Bestimmung von Korrosionsgeschwindigkeiten in flüssigen Medien mit Hilfe der Radionuklidtechnik
Authors: Vehlow J.
Published: May 2005

Die beschriebene Methode ist einfach und liefert auch bei niedrigen Korrosionsgeschwindigkeiten schon nach kurzer Zeit Ergebnisse. Die geringsten gemessenen Korrosionsgeschwindigkeiten lagen im Bereich zwischen 1 und 10 mg/m2. Tag. Diese Empfindlichkeit kann mit Hilfe von geeigneten Meßanordnungen noch weiter erhöht werden. Im Falle der Stähle kann man ohne Schwierigkeit bis zu drei Bestandteile getrennt messen, so daß auch die Untersuchung von Korrosionsvorgängen im Passiv‐ und im Übergangsbereich möglich ist. Grundsätzlich sind die γ‐ und die β‐Strahlung geeignet, die erste ist jedoch vorzuziehen, besonders zur Analyse mehrerer Bestandteile. Die Halbwertszeit der Radionuklide muß in der Größenordnung von mehreren Stunden liegen und die γ‐Energie darf nicht weniger als 100 keV betragen. Die Versuche wurden speziell mit den folgenden Systemen durchgeführt: CrNiMo‐Stähle in Schwefel‐ und Salzsäure sowie in FeCl3‐Lösung.
A Contribution to the Study of the Critical Pitting Potential of Oxide Covered Aluminum in aqueous chloride solutions
Authors: Foroulis Z. A. , Thubrikar M. J.
Published: May 2005

The critical pitting potential of oxide covered aluminum electrodes in aqueous chloride solutions has been investigated as a function of chloride ion concentration, temperature, solution pH and oxide film thickness. The steady state critical pitting potential decreases with chloride ion concentration and increasing solution temperature. Solution pH in the range 5 to 9 has no effect on the breakdown potential. The role of the oxide film thickness is to slightly increase the critical potential for passivity breakdown and intiation of pitting. It is postulated that at the critical pitting potential, passivity breakdown occurs by a process of field assisted Cl adsorption on the hydrated oxide surface and formation of a soluble basic chloride salt with a lattice cation which readily goes in solution. This process of localized dissolution of the hydrated oxide film via formation of a soluble basic aluminum chloride salt, once initiated, is likely to continue in an “autocatalytic” fashion until the oxide is locally “penetrated” and dissolution of the substrate metal begins.
Polarisationsmessungen an niedriglegierten Stählen in konzentrieten Nitratlösungen
Authors: Büuning A. , Wu Y. , Dahl W. , Schwenk W.
Published: May 2005

An sechs verschiedenen niedriglegierten Stählen wurden in Ca(NO3)2‐ und NaNO3‐Lösungen die J(U)‐Kurven mit Hilfe potentiostatischer Wechsel‐ und Halteversuche gemessen. Die instationären Kurven sind stark von der Polarisationsroutine abhängig und kennzeichnen nicht das anodische Verhalten der Werkstoffe. Die Messungen in Ca(NO3)2‐Lösungen zeigen keine Korrelation mit dem Verhalten gegenüber Spannungsrißkorrosion.

In den NaNO3‐Lösungen sind die Verhältnisse ähnlich. Im Gegensatz zu den Ca(NO3)2‐Lösungen kann aber bei mittleren Potentialwerten um UH = 0,5 V ein Beständigkeitsfeld für interkristalline Korrosionsarten vorliegen, das durch relativ hohe stationäre Stromdichten gekennzeichnet ist. Bei den Werkstoffen, die dieses Beständigkeitsfeld nicht aufweisen, sind die stationären Stromdichten deutlich geringer.

Die elektrochemischen Eigenschaften des Systems Stahl/Nitrallösungen sind äußerst kompliziert, wobei das Ruhepotential im Bereich eines Durchbruchspotentials zwischen passiver und transpassiver Korrosion liegt. Bei anodischer Polarisation können Bereiche für Anfälligkeit und Beständigkeit gegen interkristalline Korrosionsarten abwechseln.
Surfaces for interfacing protein gel electrophoresis directly with mass spectrometry
Authors: Vestling Martha M. , Fenselau Catherine
Published: May 2005

Molecular biologists and biochemists often transfer proteins separated by gel electrophoresis to membranes, such as PVDF (polyvinylidene difluoride) and nitrocellulose, for further characterization. MALDI‐MS (matrix‐assisted laser desorption/ionization mass spectrometry) can analyze proteins and peptides on a wide variety of surfaces. This review surveys the suitability of membranes and surfaces used by molecular biologists and biochemists as mass spectrometric surfaces. Matrix application considerations are reviewed. The identification of proteins by comparing the masses of peptides generated by on‐membrane digestion of proteins with theoretical digestion of database proteins is discussed. The prospects for mass spectrometric sequencing of individual peptides on membranes are presented. © 1996 John Wiley & Sons, Inc.
Particle‐induced desorption in mass spectrometry. Part I. Mechanisms and processes
Authors: Demirev Plamen A.
Published: May 2005

Desorption of (predominantly) organic molecules, induced by particles with keV to MeV energy incident on a condensed phase target, is the topic of this review. The focus is on MeV atomic ion‐induced desorption–plasma desorption (PD). Parallels with keV particle desorption–secondary ion mass spectrometry (SIMS) and fast atom bombardment (FAB)–will be presented more schematically and chiefly for comparison purposes. Current understanding of desorption mechanisms by referring to insights from theory of the processes is surveyed, and the underlying similarities as well as differences between keV and MeV particle desorption are addressed. Several connections between particle‐induced desorption and photon‐induced desorption (e.g., matrix‐assisted laser desorption/ionization‐MALDI) are briefly outlined. Results from computer simulations are used to visualize the dynamics of the desorption process. A brief description of experimental characteristics of desorbed species, aimed at elucidating the mechanisms of desorption, is given, partly to illustrate the use of mass spectrometry in fundamental studies of ion–solid interactions. Ionization pathways in PD and molecular SIMS are briejy summarized. No special emphasis is placed in describing the “generic” instrumental setup f o r particle desorption and only specific technical aspects, not treated elsewhere, eg., time and spatial correlation of the ejected species, are discussed here.

In the second part of this review (subsequent article) desorption induced by polyatomic projectiles, and effects, accompanying ion ejection, namely secondary electron emission, impact‐induced photon emission, and sputtering of clusters (fullerenes in particular), are also reviewed. In conclusion, selected examples for analytical applications of desorption methods that may illustrate current and emerging trends and perspectives are discussed. © 1996 John Wiley & Sons, Inc.
Moderne Technik des Plasmaschmelzschneidens
Authors: Born Klaus
Published: May 2005

Nach Erläuterung des Verfahrensprinzips wird die neuzeitliche Technik des Plasmaschneidens mit Argon‐Wasserstoff‐Gemischen und Spitzenelektroden beschrieben. Entsprechende Geräte, Brenner und Führungseinrichtungen werden vorgestellt. Die Zusammenhänge zwischen Anlagenleistung, Vorschubgeschwindigkeit und Blechdicke sowie Schnittqualität werden demonstriert. Ein Vergleich zu herkömmlichen thermischen Trennverfahren und ein Blick auf die Einsatzgebiete der Plasma‐Schneidtechnik schließen den Artikel ab.
Zur Tragfähigkeit schwingend beanspruchter Hochtemperatur‐Lötverbindungen
Authors: Steffens H.‐D. , Lange H.
Published: May 2005

Nach der Auswertung des einschlägigen Schrifttums werden systematische und statistisch abgesicherte Schwinguntersuchungen im Zeitfestigkeitsbereich an im Hochvakuum induktiv, jeweils mit AuNi 18 und PdNi 40 gelöteten NiCr 20 TiAl‐ und X 8 CrCo‐ NiMo 10 6‐Proben durchgeführt. Als Versuchsparameter gelten dabei nach unterschiedlicher mechanischer Lötflächenvorbereitung die Wärmebehandlung nach dem Löten sowie die Prüftemperatur. Anhand von Gefügebildern werden ergänzend häufig vorkommende Rißverläufe wiedergegeben.
Bestimmung von Reflektorgrößen mit rechteckigen Ultraschallschwingern (b/a = 0,5)
Authors: Schlengermann Udo
Published: May 2005

Bei der manuellen oder automatischen Ultraschallprüfung von voluminösen Teilen müssen statt der bekannten kreisförmigen Schallschwinger oft rechteckige Strahler eingesetzt werden. Für diese fehlen aber bisher die für Kreisschwinger gebräuchlichen Hilfsmittel zur Bestimmung von Reflektorgrößen. Deshalb werden hier theoretische und experimentelle Ergebnisse zur Bestimmung von Reflektorgrößen mit einem Rechteckschwinger vom Seitenverhältnis 2b/2a = 0,5 vorgestellt.
Quantitative holographische Prüfungen an faserverstärkten Kunststoffbauteilen aus der Raumfahrttechnik
Authors: Grünewald K. , Fritzsch W. , Wachutka H.
Published: May 2005

Die quantitative Auswertung holographischer Interferogramme erlaubt eine exakte Bestimmung von Verschiebungen und Verformungen an beliebig geformten Objekten. Die Messungen im Bereich von ca. 0 bis 100 μm können dabei mit einer Genauigkeit von ± 0,05 μm durchgeführt werden. Es wird über die Anwendung dieses Verfahrens auf einige Bauteile aus der Raumfahrttechnik berichtet, die einer thermischen bzw. mechanischen Last ausgesetzt wurden.
Zerstörungsfreie Prüfung von Tiefspaltschweißnähten mittels Durchstrahlung und Ultraschall
Authors: Steffens H.‐D. , Henderson I. D. , Echterhoff U.
Published: May 2005

Der zunehmende Bedarf an Energie erfordert Kernkraftanlagen mit immer größeren Abmessungen für die Gefäße der Druck‐ und Siedewasserreaktoren. Die Wanddicken dieser Gefäße sind auf über 200 mm angestiegen.

Große Werkstückgewichte bei extremen Einpaßgenauigkeiten der Werkstücke erfordern wirtschaftliche und leistungsfähige Schweißverfahren, die weitestgehend mechanisiert sein sollten.

Diese Anforderungen werden vom Tiefspaltschweißen erfüllt. Untersuchungen mittels konventioneller zerstörungsfreier Prüfverfahren wie Durchstrahlungs‐ und Ultraschallprüfung lassen erkennen, daß Fehlerstellen an Tiefspaltschweißungen schnell und sicher nachgewiesen werden können, da der Ort der möglichen Fehlerstellen sehr genau bekannt ist und sich eng eingrenzen läßt.

Nach Anwendung bekannter Bewertungsrichtlinien sind in den meisten Fällen unter der Voraussetzung der sicheren Beherrschung des Schweißverfahrens die Verbindungsnähte zu akzeptieren.