bluuebunny/crossref_metadata_2025_split huggingface dataset:
Myosin structure. Proximity measurements by fluorescence energy transfer
Authors: Haugland Richard P. 
Published: May 2005

AbstractThe results of energy transfer experiments on the proximity of six sites on the globular head region of myosin are discussed. A large hydrophobic crevice has been detected on each myosin head which is sufficiently large to accommodate six aromatic rings simultaneously. In the crevice is located a thiol residue not involved in activation of myosin Ca2+ ATPase and a lysine residue which is specifically trinitrophenylated with 2, 4, 6‐trinitrobenzenesulfonic acid. A second sulfhydryl whose modification activates the Ca2+ ATPase is located near the hydrophobic thiol site. The tryptophan whose fluorescence is enhanced by ATP binding is sufficiently close to the thiols and lysine residue to quantitatively transfer its energy to probes at these sites. The site of myosin ATPase has been tentatively located as being near the other five sites by energy transfer to or from synthetic chromophoric substrates. Implications of these results on the possibility of determining the location of the myosin light chain and actin binding sites are discussed.
On the enthalpy of binding of ADP to heavy meromyosin
Authors: Goodno Charles C. , Swenson Charles A. 
Published: May 2005

AbstractThe binding of ADP to heavy meromyosin has been studied by microcalorimetry. Minute amounts of myokinase interfere with binding measurements, but by selection of appropriate conditions, we can estimate that the value of the apparent ΔHbinding lies between −1.0 and −3.0 kcal per mole of ADP bound (0.3 M KCl, 2 mM MgCl2, 20 mM Tris, pH 8.00, 20°C). Values of ΔHbinding reported to date are an order of magnitude larger, and we suggest that these values are artifactual results due to myokinase contamination.
Calcium regulation of cardiac myofibrillar activation: Effects of MgATP
Authors: Solaro R. John 
Published: May 2005

AbstractIn 2 mM MgATP, 0.08 ionic strength and 1 mM free Mg++ cardiac myofibrils bound 3.5 nmoles Ca/mg protein at maximal ATPase activation. Significant amounts of Ca were also bound to cardiac myosin with these same conditions. By subtraction of this myosin‐bound Ca we obtained an estimate of 4 moles Ca bound per mole of myofibrillar troponin at maximal ATPase. We found, however, that Ca activation of myofibrillar ATPase could be estimated assuming that only two of troponin's Ca‐binding sites are engaged in regulation of crossbridge activity. Increase in MgATP from 0.3 to 5.0 mM raised the free Ca, giving half‐maximal isometric tension or ATPase. Although part of this shift is most probably due to changes in the number of rigor (nucleotidefree) actin‐myosin linkages, the rightward shift of the free Ca++‐activation relation with increase in MgATP from 2 to 5 mM appears to be due to effects of active (nucleotide‐containing) actin‐myosin linkages.
The use of aryl azido ATP analogs as photoaffinity labels for myosin ATPase
Authors: Jeng Stella J. , Guillory Richard J. 
Published: May 2005

AbstractThe synthesis of ATP analogs containing a photoactive aryl azido grouping coupled to the 3′ hydroxyl of ATP is described. The potential effectiveness of these analogs in the investigation of nucleotide‐binding regions is outlined and this effectiveness demonstrated by their photodependent inhibition of subfragment 1 ATPase. The use of 14C‐labeled azido ATP demonstrates an almost stoichiometric covalent binding of the analog.Because of their potential application to other systems, a number of reactions describing the reactivity of the 3′ hydroxyl of the nucleotide ribose are outlined in an Appendix.
Synthesis of adenosine triphosphate by way of potassium‐sensitive phosphoenzyme of sodium, potassium adenosine triphosphatase
Authors: Post Robert L. , Toda Gotaro , Kume Shoji , Taniguchi Kazuya 
Published: May 2005

AbstractThe sodium and potassium ion pump is an intrinsic enzyme of plasma membranes. In these experiments it was driven backward in a transient two‐step operation involving, first, phosphorylation of the enzyme from inorganic phosphate, and second, transfer of the phosphate group from the enzyme to ADP upon addition of a high concentration of Na+. There was no evidence of a significant concentration gradient across the membranes. Na+ presumably reached the solutions on both faces of the membrane simultaneously and provided the energy for synthesis simply as a consequence of ligand binding. An interaction free energy between the free energy of the binding of Na+ presumably reached the solutions on both faces of the membrane simultaneously and provided the energy for synthesis simply as a consequence of ligand binding. An interaction free energy between the free energy of the binding of Na+ and the free energy of hydrolysis of the phosphate group on the enzyme was estimated. The experiments also suggested a feature of the transport mechanism. This is control by phosphorylation of access pathways from the solutions in contact with the faces of the membrane to an active center for cation binding. In the dephosphoenzyme access would be to the intracellular solution and in the phosphoenzyme access would be to the extracellular solution.
Nonrandom distribution of receptors for melanocyte‐stimulating hormone on the surface of mouse melanoma cells
Authors: Varga Janos M. , Saper Mark A. , Lerner Aaron B. , Fritsch Peter 
Published: May 2005

AbstractAn improved bubble method was developed for applying an ultrathin layer of nuclear track emulsion on the surface of cells labeled with I125‐MSH. The autoradiographs of I125‐MSH binding indicate a nonrandom distribution of receptors on the surface of mouse melanoma cells. It is suggested that MSH receptors are displayed in clusters previous to an independently of their exposure to the hormone.
Concanavalin a perturbation of membrane enzymes of mammary gland
Authors: Carraway Coralie A. , Carraway Kermit L. 
Published: May 2005

AbstractThe plant lectin concanavalin A (Con A) specifically inactivates the 5′ ‐nucleotidase of a plasma membrane‐enriched fraction from lactating mammary gland. The lectin also causes an activation of the membrane Mg++ ‐ATPase, but does not affect galactosyltransferase or alkaline phosphatase. The enzyme perturbations are prevented by α‐methylmannoside, an inhibitor of Con A binding, indicating that specific binding to carbohydrate structures rather than nonspecific protein‐protein interaction is involved. Solubilization of the 5′ ‐nucleotidase in detergents (0.2% Triton X‐100 or 1% deoxycholate) does not prevent Con A inactivation, indicating that incorporation into the membrane structure is not a requirement for the Con A effect. The results suggest that Con A inactivates the 5′ ‐nucleotidase by a direct interaction with the enzyme and that this enzyme is a Con A receptor site on the surface of mammary cells.
Selective modification of cell surface proteins and thymidine transport in hamster cells exposed to cholera toxin
Authors: Rieber M. , Bacalao J. 
Published: May 2005

AbstractThe increased adherence and morphological response which occurs in Chinese hamster ovary cells as a result of exposure to cholera toxin is paralleled by modification in the relative exposure of outer proteins. Mild proteolysis treatment of the cells prelabeled with [3H] glucosamine reveals a markedly different kinetics of release of external glycopeptides as a result of exposure to cholera toxin. Selective alterations in external tyrosyl‐rich proteins can also be detected by lactoperoxidase‐catalyzed radioiodination. The above modifications are accompanied by a decrease in the rate of thymidine uptake by toxintreated cells.
Proteins of the hepatoma tissue culture cell plasma membrane
Authors: Tweto John , Friedman Else , Doyle Darrell 
Published: May 2005

AbstractThe specificity of lactoperoxidase‐catalyzed iodination for the proteins of the hepatoma tissue culture cell plasma membrane was examined by histochemical, biochemical, and cell fractionation techniques. Light microscope autoradiography of sectioned cells shows the incorporated label to be localized primarily at the periphery of the cell. Most of this label can be released from the cell by trypsin but not by collagenase or hyaluronidase. The label is recovered from the cells as either monoiodotyrosine or diiodotyrosine after hydrolysis of cell extracts with a mixture of proteolytic enzymes. The label co‐purifies during cell fractionation with an authentic liver cell plasma membrane marker enzyme, 5′‐nucleotidase. Thus, the incorporated iodide is itself a valid marker for those membrane polypeptides having tyrosine residues accessible to the lactoperoxidase. The polypeptide complexity of the purified plasma membrane was examined by high resolution dodecyl sulfate‐polyacrylamide gel electrophoresis. At least 50 polypeptides in the membrane are accessible to iodination. These polypeptides probably represent the bulk of the protein mass of the membrane and iodinating them does not affect cell viability, growth rate, or cell function. Labeling experiments with fucose and glucosamine show that at least nine of the iodinated peptides may be glycoproteins.
Heterogeneity in the conformation of different protein fractions from the human erythrocyte membrane
Authors: Holzwarth G. , Yu John , Steck Theodore L. 
Published: May 2005

AbstractWe have isolated 5 families of proteins from human red blood cell membranes and characterized their secondary structure by ultraviolet circular dichroism measurements. The protein families were prepared by selective solubilization from ghosts under nondenaturing conditions. We find that the intact ghost has a mean α‐helix fraction of 0.37, whereas a low‐ionic‐strength extract (bands 1, 2, 5, “spectrin”) has a substantially higher helix fraction, 0.55. Further extraction of the ghosts with para‐chloromercuribenzoate yields bands 2.1, 4.1, 4.2, and 6; their helix content is only 0.17. Finally, the major intrinsic protein, band 3, was solubilized by a nonionic detergent. Its helix fraction is 0.38.
DNA synthesis in cultured human fibroblasts: Regulation by 3′ : 5′‐cyclic amp
Authors: Rechler Matthew M. , Bruni Carmelo B. , Podskalny Judith M. , Carchman Richard A. 
Published: May 2005

AbstractThe addition of serum to density‐inhibited human fibroblast cultures induced a wave of DNA synthesis, measured as [3H] thymidine incorporation into acid‐precipitable material, beginning after 8–12 hr and reaching maximum levels at 16–24 hr. Addition of dibutyryl‐3′ : 5′‐cyclic AMP (DBcAMP) together with serum inhibited [3H] thymidine incorporation by 75–95%. When DBcAMP was added for the first 4 hr of serum stimulation and then removed, the wave of DNA synthesis was not delayed. This suggested that serum could induce DNA synthesis even though cyclic AMP concentrations were maintained at high levels by DBcAMP during this initial period. These results are inconsistent with the hypothesis that it is the immediate transient reduction in 3′ : 5′‐cyclic AMP concentration following the addition of serum that triggers DNA synthesis. By contrast, DBcAMP added 8 hr after serum inhibited [3H] thymidine incorporation to the same extent as DBcAMP added at the same time as serum. This indicated that a step essential for DNA synthesis and occurring late in G1 was inhibited by high concentrations of 3′ : 5′‐cyclic AMP.
Some kinetic and chromatographic properties of detergent‐dispersed adenylate cyclase
Authors: Johnson Roger A. , Garbers David L. , Pilkis Simon J. 
Published: May 2005

AbstractStudies on the reaction kinetics and chromatographic properties of detergent‐dispersed adenylate cyclase are described. Detergent‐dispersed enzyme was prepared from whole rat cerebellum and from partially purified plasma membranes from rat liver.Data were simulated to fit kinetic models for which an inhibitor is added in constant proportion to the variable substrate. Models were chosen to distinguish whether the adenylate cyclase reaction may be controlled by an inhibitory action of free ATP−4 (or HATP−3) or by a stimulatory action of free divalent cations. The various kinetic models were then tested with the dispersed brain adenylate cyclase with both Mg++ and Mn++ and in two different buffer systems. The experimental data indicate that this enzyme has a distinct cation binding site, but exhibits no significant inhibition by HATP−3 or ATP−4.The detergent‐dispersed adenylate cyclase both from liver plasma membranes and from brain have been chromatographed on anion exchange material and have been chromatographed on anion exchange material and have been subjected to gel filtration. The presence of detergent was required for elution of cyclase activity from DEAE‐Sephadex but was not required when DEAE‐agarose was used. Dispersed brain cyclase was also chromatographed on agarose‐NH(CH2)3 NH(CH2)3‐NH2 which exhibits both ionic and hydrophobic properties. Fifty percent of the applied activity was recovered with a fivefold increase in specific activity. The data suggest that the relative effectiveness of a given chromatographic procedure for detergent‐dispersed adenylate cyclase may reflect the in fluence of both hydrophobic and ionic factors.
Studies of bacterial chemotaxis in defined concentration gradients. A model for chemotaxis toward L‐serine
Authors: Dahlquist F. W. , Elwell R. A. , Lovely Peter S. 
Published: May 2005

AbstractThe details of the chemotactic response of Salmonella typhimurium to gradients of L‐serine have been examined in some detail. Two relatively macroscopic techniques have been employed to measure the bacterial response. These include measurements of the average velocity as the bacterial population moves toward attractants, and measurement of the upward‐to‐downward flux ratio, R, in the stable preformed attractant gradients. The dependence of the average velocity on gradient appears to be hyperbolic in nature, while the flux ratio depends linearly on the gradient. These data suggest a microscopic model for the dependence of bacterial behavior on the serine gradient. The model involves a linear dependence of the mean lifetime of a bacterial trajectory on the gradient for those bacteria moving toward higher attractant concentration. Those moving toward low concentrations of attractant do not change the mean duration of their trajectories, or the speed at which a given bacterium swims through the solution. This model generates the observed dependences of the average velocity and flux ratio on gradient. Interpretation of the experimental data suggests that a gradient which increases serine concentration by a factor of 2 in 10 mm is sufficient to double the average duration of a trajectory for a bacterium moving directly up the gradient. The concentration dependence of the chemotactic response to serine is more complicated. It suggests that more than one receptor of serine may be involved in determining chemotactic behavior to this attractant.
Immunological studies of acetylcholine receptors
Authors: Lindstrom Jon 
Published: May 2005

AbstractImmunochemical techniques for the study of acetylcholine receptors are described. Immunization of rabbits, rats, guinea pigs, and goats with acetylcholine receptor protein purified from Electrophorus electric organ tissue results in muscular weakness and death due to impaired neuromuscular transmission. Serum from immunized animals contains high concentrations of antibodies directed at receptors from the electric organ and low concentrations of antibodies directed at receptors from skeletal muscle. The detailed similarities between the disease of receptor‐immunized animals, “experimental autoimmune myasthenia gravis” (EAMG), and myasthenia gravis are compared. Reactions of antisera from animal with EAMG with receptor from Electrophorus and Torpedo are studied. Antireceptor antibodies in these antisera are directed predominantly at determinants other than the acetylcholine‐binding site.
Reconstitution of a purified acetylcholine receptor
Authors: Michaelson D. M. , Duguid J. R. , Miller D. L. , Raftery M. A. 
Published: May 2005

AbstractDialysis of the purified acetylcholine receptor from Torpedo californica electroplax with lipids from the same organ results in a vesicular membrane system in which the receptor is embedded in the bilayer and oriented so that most of the neurotoxin‐binding sites appear to be on the outer surface. The constituted vesicles are chemically excitable by acetylcholine and carbamylcholine, as measured by 22Na+ efflux. The excitability is specifically blocked by the antagonist α‐bungarotoxin. These results demonstrate that the purified reconstituted receptor system not only can specifically bind neurotransmitter but also can trigger ion translocation. It therefore has the properties necessary to effect postsynaptic depolarization in vivo.
The role of cell membrane in the antiviral effect of interferon
Authors: Pitha Paula M. , Vengris Vitolis E. , Reynolds Fred H. 
Published: May 2005

AbstractThe mechanism of interferon action in human fibroblasts has been studied by use of both antisera to human fibroblast interferon and the antisera to the surface of human fibroblast cell. The anti‐interferon serum completely neutralized the antiviral effect of human fibroblast interferon. Interferon antiserum prevented the intracellular antiviral state from developing when added to the medium of the cells in which interferon synthesis had already been induced by poly (I · C). This suggests that development of the antiviral state involves interferon interaction with the external part of the producing cell. Treatment with the serum directed against the surface of human fibroblast cells failed to inhibit the antiviral activity of human interferon in these cells.In addition, the effect of gangliosides on the antiviral activity of human interferon was studied and it was found that human interferon binds to gangliosides and that this interaction leads to inactivation of the antiviral effect of interferon. Pretreatment of human fibroblasts with gangliosides had no effect on the sensitivity of these cells to exogenous interferon.
Effect of polyamines on the uptake of poly (2′‐fluoro‐2′‐deoxyuridylic acid) by a mammalian cell line
Authors: Janik Borek , Sommer Ronald G. 
Published: May 2005

AbstractVERO cells can take up poly(dUfl)1 from the medium. The uptake involves surface adsorption and, most probably, intracellular penetration. Part of the poly(dUfl) is hydrolyzed during incubation with the cells but the hydrolysis products are not incorporated into de novo synthesized nucleic acids. The uptake is reduced by serum and stimulated by polycationic ionenes. The magnitude of stimulation depends on the structure of the ionene and the treatment regimen.
Effect of essential fatty acid deficiency on activity of liver plasma membrane enzymes in the rat
Authors: Louis S. L. , Brivio‐Haugland R. P. , Williams M. A. 
Published: May 2005

AbstractLiver plasma membranes (LPM) were isolated from rats fed an essential fatty acid‐supplemented diet (+EFA) or from rats fed an essential fatty acid‐deficient diet (‐EFA). The proportions of linoleate and arachidonate in membrane total fatty acids in the −EFA preparations were one‐half or less than the values for the +EFA preparations. Basal, F−, or glucagon‐stimulated adenylate cyclase activities were significantly lower in EFA‐deficient livers than in nondeficient ones. Addition of GTP significantly enhanced glucagon‐stimulated adenylate cyclase in both groups, but extent of stimulation above basal was greater in EFA‐deficient livers. Portal vein injection of glucagon in vivo resulted in significantly higher cAMP formation in +EFA livers than in −EFA livers. When glucagon was used in vitro at 1–1,000 nM, stimulation of adenylate cyclase remained lower in EFA‐deficient membranes, but extent of stimulation above basal activity was larger in −EFA membranes than in +EFA. Total Na+, K+ (Mg2+)‐ATPase from EFA‐depleted LPM exhibited significantly higher values of apparent Km and Vmax. 5′‐Nucleotidase activity, in contrast, was considerably decreased in EFA‐deficient rats. These findings show that, in animals, changes in unsaturated fatty acid composition can affect the properties of membrane‐bound enzymes. These alterations could be due to changes in membrane physical properties and/or prostaglandin formation.
Drug‐resistant mutants of chinese hamster ovary cells possess an altered cell surface carbohydrate component
Authors: Juliano Rudy , Ling Victor , Graves James 
Published: May 2005

AbstractSurface label experiments using the galactose oxidase‐[3 H] ‐borohydride technique reveal that cells from drug‐resistant Chinese hamster ovary clones possess a surface carbohydrate component of apparent molecular weight 165,000 which is absent from wild‐type cells. The component may also be demonstrated by [14C] glucosamine incorporation but not by [3 H] leucine incorporation or by the lactoperoxidase surface labeling reaction.
A scanning electron microscope study of concanavalin a receptors on retinal rod cells labeled with latex microspheres
Authors: Molday Robert S. 
Published: May 2005

AbstractCon A‐methacrylate microsphere conjugates prepared by a two‐step glutaraldehyde reaction were used to label Con A‐binding sites on bovine rod photoreceptor cells for visualization by scanning electron microscopy. A dense distribution of markers was observed on the surface of the rod outer segment, the inner segment, and the synaptic region. Disk membranes also appear to be heavily labeled with the Con A‐microsphere conjugates. The Con A inhibitor, α‐methyl mannoside, inhibited the binding of the conjugate to the surface of these visual cells.
Some changes in the properties of dynein ATPase in situ and after extraction following heat treatment of cilia
Authors: Blum J. J. , Hayes Alveron 
Published: May 2005

AbstractGlycerol‐extracted cilia from Tetrahymena pyriformis were demembranated by treatment with Triton X‐100 and then heated for up to 30 min at temperatures between 34–38°C. Heat treatment carsed an uncoupling of the ATPase from motility as indicated by an increase in ATPase activity and a loss of pellet height response. After heat treatment, the ATPase activity of the dynein in situ differed from that in unheated cilia as shown by an increased sensitivity to a lower temperature of assay (0°C) and by a loss of the activation normally observed upon reaction with N‐ethylmaleimide or p‐phenylenedimaleimide. Upon extraction of the heat‐treated cilia by Tris‐EDTA, there was a large loss in ATPase activity so that the heat‐treated cilia yielded a crude dynein fraction with a lower specific activity compared with that obtained from unheated controls. The difference was not due to a change in the amount of protein recovered or in the amount of ATPase activity which remained unextracted. Resolution of the crude dynein by sucrose density sedimentation indicated that activity was lost from both the 14S and 30S peaks but more so from the latter than from the former. Thus dynein in situ in cilia in which the ATPase has been uncoupled from motility by gentle heat treatment differs in several important respects from dynein inside unheated cilia.
Isolated cortical granules: A model system for studying membrane fusion and calcium‐mediated exocytosis
Authors: Vacquier Victor D. 
Published: May 2005

AbstractCortical granules are secretory vesicles bound to the inner surface of the plasma membrane of sea urchin eggs. Intact granules can be isolated by shearing away the cytoplasm of eggs which have been bonded to a protamine‐coated surface. When Ca2+ is added to preparations of isolated granules the granules fuse with each other and release their contents. It is believed that isolated cortical granules may be an excellent model system for the biochemical study of exocytosis.