bluuebunny/crossref_metadata_2025_split huggingface dataset:
Time‐dependent fluorescence depolarization and lifetime studies of myosin subfragment‐one in the presence of nucleotide and actin
Authors: Mendelson Robert , Putnam Susan , Morales Manuel 
Published: May 2005

AbstractTime‐dependent fluorescence depolarization and lifetime studies have been made on myosin subfragment 1 to obtain information about mobility changes and dye environment changes when different nucleotides are added. Data are reported for static and actively hydrolyzing systems containing G‐ and F‐actin. Preliminary data indicate that myosin labeled with the fluorophore 1,5 IAEDANS1‐and treated with DTT preserves its actin‐activated Vmax. S1 prepared in this manner gives lifetime changes which are nearly identical for all systems studied. S1 labeling without DTT addition gives a pattern of lifetimes similar, though not identical to ESR work. Either type of labeling produces no observable change in the polarization decay, and we set an upper limit of 15% length change for the elongate S1. An unusually long fluorescence decay lifetime for the S1‐Mg++ ATP‐G‐actin system is found which may indicate a new acto‐S1 state stabilized by G‐actin. The method for obtaining the bound fraction of S1's in the presence of actin is presented and applied to the S1‐F‐actin‐Mg++ ATP system. Qualitative agreement is obtained with other methods.
Studies of substructure and tightly bound nucleotide in bacterial membrane ATPase
Authors: Abrams A. , Jensen C. , Morris D. 
Published: May 2005

AbstractHighly purified preparations of Streptococcus faecalis ATPase contain a similar but inactive protein detected by prolonged polyacrylamide gel electrophoresis. The inactive protein appears to arise by proteolytic cleavage of the major subunits in the enzyme. By use of a new technique, subunit analysis in SDS gels was performed on the enzyme band and the inactive protein band excised from a polyacrylamide gel after electrophoresis. The results indicated that the ATPase has the composition α3β3γ in which α = 60,000, β = 55,000, and γ = 37,000 daltons. The inactive protein appears to have the composition (f)6 in which f = 49,000 daltons. There is also evidence that the enzyme band contains some slightly modified forms of the ATPase, such as α3β2 (f)γ. The inactive protein lacks the capacity for tight nucleotide binding.Our experiments show that the tight ATPase‐nucleotide complex formed in S. faecalis cells (the endogenous complex) behaves differently from the tight complex formed in vitro (the exogenous complex). We prepared a doubly labeled complex containing endogenous 32P‐labeled ADP and ATP and exogenous 3H‐labeled ADP. We observed that the addition of free nucelotide to the doubly labeled ATPase displaced the exogenous bound ligand from the enzyme but not the endogenous bound nucleotide. We suggest that the displaceable and nondisplaceable forms of the tight ATPase‐nucleotide complex correspond to two different conformational states of the enzyme.
Interaction of myosin and paramyosin
Authors: Epstein Henry F. , Aronow Bruce J. , Harris Harriet E. 
Published: May 2005

AbstractThe interaction of myosin and paramyosin was investigated by enzymological and ultrastructural techniques. The actin‐activated Mg+2 ATPase of rabbit skeletal muscle myosin can be inhibited by clam adductor paramyosin. Both proteins must be rapidly coprecipitated to form filaments for this inhibition. Slowly formed cofilaments are fully activatable by F‐actin. In both cases, the cofilaments possess unique structural characteristics when compared to homofilaments.The mode of inhibition appears to be competitive when different concentrations of paramyosin and F‐actin are compared. The apparent affinity of the myosin heads for actin is reduced by the presence of paramyosin within rapidly reconstituted thick filaments. These results suggest that paramyosin may serve as part of a relaxing mechanism within invertebrate muscles. It is unlikely that paramyosin plays a role in the initiation and maintenance of catch within specialized molluscan muscles.
Calcium and magnesium ATPASES of the spectrin fraction of human erythrocytes
Authors: Kirkpatrick F. H. , Woods G. M. , La Celle P. L. , Weed R. I. 
Published: May 2005

AbstractUsing a rapid method of preparation, spectrin has been isolated from human erythrocytes and its ATPase activity investigated. The ATPase activity with calcium has two distinct components, ‐one with optimal activity when calcium and ATP are of equal concentration (low‐Ca‐ATPase) and another which is activated above 1 mM CaCl2 and is maximal at 100 mM CaCl2. There is also a Mg‐ATPase with maximal activity at 10 mM MgCl2. The high‐Ca‐ATPase of spectrin, but not the low‐Ca‐ATPase, is inhibited by magnesium, while the Mg‐ATPase is inhibited by Ca in excess of ATP. None of these activities exhibits the calcium‐stimulated magnesium‐dependent activity characteristic of the red cell calcium pump.
Possible role of firmly bound ATP in the energy transduction of photosynthetic membranes
Authors: Lutz Hans U. , Beyeler Walter , Pflugshaupt Christian , Bachofen Reinhard 
Published: May 2005

AbstractChromatophores of Rhodospirillum rubrum and spinach chloroplasts contain firmly bound ATP that is rapidly labeled along with ADP in the presence of 32Pi and endogenous nucleotides. The labeling is not entirely dependent on light. In chloroplasts three types of bound ATP can be defined methodologically by their extraction properties: buffer‐soluble; acid‐soluble; and SDS‐soluble or firmly bound ATP. Extensive washing of the chloroplasts does reduce buffer‐soluble but not acidsoluble and firmly bound APT. Buffer‐soluble [32P] ATP is almost exclusively γ labeled while acid‐soluble and firmly bound ATP are labeled in the β and γ position equally. CCCP, desaspidin, and phlorizin do not inhibit the labeling of firmly bound ATP, whereas the phosphorylation is almost abolished. However, EDTA and NEM pretreatments of the choloroplasts affect both reactions similarly.The postillumination [32P] ATP synthesis with chromatophores can be inhibited by adding ATP to the incubation mixture after illumination if 32Pi is included only during the dark incubation, but is without effect if 32Pi is present only during illumination. On the other hand, ADP added after illumination inhibits post‐illumination [32P] ATP formation in both chromatophores and chloroplasts only if 32Pi is present during illumination. The data can be explained by a coupling factor having two sites, as proposed previously on the basis that firmly bound ATP does not transfer its phosphoryl group but seems to drive a synthesis of acid‐soluble ATP which incorporates free phosphate (Lutz, H. U., Dahl, J. S., and Bachofen, R.), Biochim. Biophys. Acta 347:359 [1974].
ATP‐stimulated transmitter release and cyclic amp synthesis in isolated chromaffin granules
Authors: Hoffman Philip G. , Zinder Oren , Nikodijevic Olga , Pollard Harvey B. 
Published: May 2005

AbstractATP stimulates chromaffin granules from the bovine adrenal medulla to release epinephrine and specific soluble proteins. ATP analogs substituted in the β‐γ‐position with either nitrogen or carbon were also found to be effective at inducing release from isolated chromaffin granules. However, an ATP analog substituted at the α‐β position with carbon was strongly inhibitory. Cyclic AMP was also found to be synthesized by isolated chromaffin granules under release conditions. ATP analogs were effective as substrates for adenylate cyclase in the same order as their efficiency for inducing release from vesicles. Hydrolysis at the β‐γ linkage of ATP therefore is probably not necessary for release; however, hydrolysis at the α‐β position may be important in the release process. Cyclic AMP may be produced and play a regulatory role in this event.
Cooperative properties of hormone receptors in cell membranes
Authors: Meyts Pierre De 
Published: May 2005

AbstractThe binding of many polypeptide hormones to cell surface receptors does not appear to follow the law of mass action. While steady–state binding data are consistent in many cases with either heterogeneous populations of binding sites or interactions of the type known as negative cooperativity, study of the kinetics of dissociation of the hormone receptor complex allows an unambiguous demonstration of cooperative interactions. Negative cooperativity, which seems to be wide‐spread among hormone receptors, provides exquisite sensitivity of the cell at low hormone concentrations while buffering against acutely elevated hormone levels. The molecular mechanisms underlying the cooperativity are still largely unknown. Cooperativity may stem from a conformational transition in individual receptors or involve receptor aggregation in the fluid membrane (clustering) or more extensive membrane phenomena. Thus, new models of hormone action must be considered which integrate the progress in our knowledge of both the complex mechanisms regulating hormone binding to their surface receptors, and the dynamic properties of the cell membrane.
Preliminary characterization of the acetylcholine receptor in human erythrocytes
Authors: Huestis Wray H. 
Published: May 2005

AbstractThe response of human erythrocytes to cholinergic ligands was studied with an electron spin resonance assay. The membrane response to carbamyl choline was found to be antagonized by atropine and, in the absence of calcium, by tetrodotoxin. Experiments with resealed ghosts showed that the membrane response to carbamyl choline required ATP and calcium. Reductive alkylation of intact cells eliminated the cholinergic response, but the presence of saturating amounts of carbamyl choline protected the putative receptor against inactivation. Affinity labeling was used to demonstrate an apparent molecular weight of 41,000 for the carbamyl choline‐binding species. A lipid vesicle extraction technique was used to induce a specific cation permeability defect in intact cells. Preliminary investigation of this phenomenon is described.
Light‐Induced tetracycline accumulation by rhodopseudomonas sphaeroides
Authors: Weckesser Jürgen , Magnuson James A. 
Published: May 2005

AbstractLight has been used as a primary energy source in studies of tetracycline transport by Rhodopseudomonas sphaeroides. Accumulation of the antibiotic occurs in light, while efflux occurs in dark. Both fluorescence enhancement and radioisotopic tracing have been used to monitor transport. Km's obtained from both techniques are similar. Light‐induced accumulation of tetracyclines is inhibited by a variety of inhibitors, including antimycin A, N‐ethylmaleimide, carbonylcyanide m‐chloro‐phenylhydrazone, and 2,4‐dinitrophenol. A rapid efflux is observed after loading when cells are placed in the dark or treated with inhibitors.
The surface events at fertilization: The movements of the spermatozoon through the sea urchin egg surface and the roles of the surface layers
Authors: Schatten Gerald , Mazia Daniel 
Published: May 2005

AbstractThe sea urchin egg surface at fertilization has been examined with the scanning electron microscope to reveal the movements of the spermatozoon from the exterior, through the surface layer, and into the egg cytoplasm. The layers that the spermatozoon encounter have been studied to determine their physical and chemical natures and their role in early development.By studying the outside of whole eggs and the inner face of surfaces isolated shorlty after fertlization, it has been possible to complie data on the movements of the spermatozoon through the egg surface. The spermatozoon initially contacts the egg with the elongated acrosomal process. The vitelline sheet, the outermost layer of the egg, separates slightly next to the attached spermatozoon. As membrane from the egg engulfs the spermhead, the cortical granules start to discharge their contents, and a spreading surface deformation, concommitant with a distortion of the fibrous cortex, is initiated. A cluster of elongate microvilli surround the perpendicularly fusing spermatozoon. These microvilli interdigitate as the spermatozoon is forced to lie upon the egg surface between the plasma membrane and the matrix of cortical fibers. The spermatozoon the rotates additionally to enter the egg cytoplasm with the posterior end first; it has rotated 180° through the cell surface. Finally, it detaches into the egg cytoplasm, leaving a scar in the cortex through which it penetrated.The egg cortex, previously unobserved by electron microscopy, is revealed to be comosed of 50–200 nm fibers. At fertilization they are uniformly organized but during later development this order is lost. The cortex is from 0.2–0.5 μm thick and is a contractile structure.The role of the outer surface in releasing the cell from the metabolic constraints of the unfertilized egg is shown, and the apparent in the mobilities of the membranes derived from the sperm and the egg are demonstrated. The relation of these layers to the movements of the speratozoon, to the activation of the egg, to the block to polyspermy, and to each other are discussed.
Interactions of neurotoxins with the action potential Na+ ionophore
Authors: Catterall William A. , Ray Radharaman 
Published: May 2005

AbstractFour neurotoxins that activate the action potential Na+ ionophore of electrically excitable neuroblastoma cells interact with two distinct classes of sites, one specific for the alkaloids veratridine, batrachotoxin, and aconitine, and the second specific for scorpion toxin. Positive heterotropic cooperativity is observed between toxins bound at these two classes of sites. Tetrodotoxin, a specific inhibitor of the action potential Na+ current, inhibits activation by each of these toxins in a noncompetitive manner (KI = 4–8 nM). These results suggest the existence of three functionally separable components of the action potential Na+ ionophore: two regulatory components, which bind activating neurotoxins and interact allosterically in controlling the activity of a third ion‐transport component, which binds tetrodotoxin. The dissociation constant for scorpion toxin binding is increased 10‐fold by depolarization of the cells with K+, suggesting that the scorpion toxin binding site is located on a voltage‐sensitive regulatory component of the ionophore.
The gating currents of sodium channels: Pore‐population‐size effects
Authors: Rayner M. D. , D'Arrigo J. S. , Conquest L. L. 
Published: May 2005

AbstractSodium‐channel behavior has been modeled in order to determine the answer to the following question: How large must a population of “on‐off” Sodium pores be before the inherently random behavior of the individual channels becomes smoothed to yield the expected gating current‐conductance relationships which would be predicted from an infinite pore array? Results of this analysis show that for the “opening” situation, an excellent fit was obtained whenever more than about 10 pores were considered. Significant discrepanciesd were observed in the “Closeing” situation, however, for pore arrays of 50 or less. Marked hysteresis is apparent in the behavior of small pore populations.
Structure and control of assembly of cytoplasmic microtubules in normal and transformed cells
Authors: Fuller G. M. , Brinkley B. R. 
Published: May 2005

AbstractIndirect immunoflurescence analyses using antibodieis directed against 6S tubulin have shown an elaborate cytoplasmic microtubule complex (CMTC) in nontransformed cells in culture. The CMTC is strikingly altered in cells that have been transformed spontaneously by viruses or by chemicals. Assembly of microtubules in vitro and in vivo is markedly inhibited in the presence of elevated levels of calcium. Alteration of the surface of normal cells by brief treatment with low concentrations of trypsin initiate a rapid breakdown of cytoplasmic microtubules. Finally, a hypothesis is presented relating microtubule assembly and surface membrane modulation suggesting that calcium is the primary modulating signal.
Effects of RNase and RNA on in vitro aster assembly
Authors: Zackroff Robert V. , Rosenfeld Arline C. , Weisenberg Richard C. 
Published: May 2005

AbstractRNase alters the in vitro assembly of spindle asters in homogenates of meiotically dividing surf clam (Spisula solidissima) oocytes. Some effects of RNase, such as reduced astral fiber length, appear nonenzymatic and probably result from RNase binding to tubulin. However, RNase‐induced changes in the microtubule organizing center are also observed. Since other polycations can mimic RNase effects, the existence of an RNA component of the spindle organizing center remains uncertain. Effects of RNase and other polycations on astral fiber length can be prevented and reversed by the RNase inhibitor, polyguanylic acid. Polyguanylic acid can also augment astral fiber length in the absence of added RNase or other polycations. Augmentation by polyguanylic acid is favored by high ionic strength, and can be duplicated by polyuridylic acid and, with less efficiency, by polyadenylic acid. Polucytidylic acid and unfractionated yeast RNA, however, are unable to augment aster assembly. Polyguanylic acid can also augment the length of astral fibers on complete spindles isolated under polymerizing condition. These results demonstrate that specfic polyribonucleotides can alter spindle assembly in vitro. The presence of an inhibitor of microtubule assembly in Spisula oocytes, which can be inactivated by specific RNAs, is suggested.
Energy sources for amino acid transport in animal cells
Authors: Heinz Erich , Geck Peter , Pietrzyk Christian , Burkhardt Gerhard , Pfeiffer Barbara 
Published: May 2005

AbstractThe existence of an electrogenic Na+ pump in Ehrlich cells which substantially contributes to the membrane potential, previously derived from the distribution of the lipid soluble cation tetraphenylphosphonium (TPP+), could be confirmed by an independent method based on the quenching of fluorescence of a cyanine dye derivative, after the mitochondrial respiration had been suppressed by appropriate inhibitors. The mitochondrial membrane potential, by adding to the overall potential as measured in this way is likely to cause an overestimation of the membrane potential difference (p.d.). But since this error tends to diminish with increasing pump activity, the true p.d. of the plasma membrane should easily account for the driving force to drive the active accumulation of amino acids in the absence of an adequate Na+ concentration gradient. Accordingly, the F2‐aminoisobutyric acid (AIB) uptake rises linearly with the distribution of TPP+ at constant Na+ concentrations, suggesting that each responds directly to membrane potential. There is evidence that the electrogenic (free) movement of Cl− is slow, at least at normal p.d., whereas a major part of the Cl− movement across the cellular membrane appears to occur by an electrically silent Cl−‐base exchange mechanism. By such a mode Cl−, together with an almost stoichiometric amount of K+, may under certain conditions move into the cell against a high adverse electrical potential difference. This “paradoxical” movement of K+Cl− contributing to the deviation of the Cl− distribution from the electrochemical equilibrium distribution, is not completely understood. It is insensitive towards ouabain but can almost specifically be inhibited by furosemide. As a likely explanation a H+–K+ exchange pump was previously offered, even though unequivocal evidence of such a pump is so far lacking. According to available evidence the electrogenic movement of free Cl− is too small, at least at normal orientation of the p.d., to significantly shunt the electrogenic pump potential so that the establishment of such a potential is plausible. The evidence presented is considered strong in favor of the gradient hypothesis since even in the absence of an adequate Na+ concentration gradient, the electrogenic Na+ pump will contribute sufficient extra driving force to actively transport amino acid into the cells.
Perspectives and limitations of resolutions–reconstitution experiments
Authors: Racker Efraim 
Published: May 2005

AbstractReconstitutions 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

AbstractPlants 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.