bluuebunny/crossref_metadata_2025_split huggingface dataset:
Crosslinking studies on the CA2+, MG2+‐activated ATPase of escherichia coli
Authors: Bragg Philip D. 
Published: May 2005

AbstractCrosslinking of membrane proteins of Escherichia coli with dithiobis (succinimidyl propionate) (DSP) resulted in loss of several enzyme activities including the Ca2+, Mg2+‐activated ATPase. This enzyme was crosslinked by DSP to the membrane and was not released by dialysis at low ionic strength in the absence of dithiothreitol which could cleave the crosslinking group. DSP inactivated both phosphohydrolase and coupling activities of the solubilized ATPase. Loss of hydrolytic activity could be correlated with the extent of reaction of the α and/or β subunits of the enzyme. The loss of coupling activity appeared to be associated with modification of the γ and/or δ subunits.
Chemical and conformational events in regions of the myosin
Authors: Szent‐Györgyi Andrew G. 
Published: May 2005

AbstractStructural changes in myosin may take place within the heavy chains, or may represent rearrangement of subunits. Changes induced by ATP and possibly restricted to the heavy chains are indicated by several different optical studies, including changes in absorbence of aromatic residues (1), changes in the fluorescene of dyes coupled to myosin (2), changes in electron spin resonance spectrum (3), and changes in the intrinsic fluorescence of tryptophan residues (4, 5). A calcium dependent subunit rearrangement in scallop myosin is indicated by fluorescene depolarization studies (Cheung and Szent‐Györgyi, unpublished observations). In scallop muscles, calcium regulates contraction by directly reacting with myosin (6), and the changes in the quaternary structure of myosin are likely to be related in the way calcium control works.At first I will disucss the changes of fluorenscene that Drs. Weber, Fasman, and I have described several years ago (4). Then I will speculate on the implications of a subunit rearrangement on the mechanism of regulatory light chain funciton.
Does myosin‐substrate interaction in vitro result in a delocalized conformation change?
Authors: Cassim Joseph Y. , Lin Tsung‐I 
Published: May 2005

AbstractThe effect of substrate on the far UV (185–250 nm) and near UV (250–325 nm) circular dichroism (CD) of myosin and heavy meromyosin (HMM) was studied. The following results were obtained with the addition of ATP (during various conditions of hydrolysis), ADP, and pyrophosphate: (1) no changes were observed in the far UV CD, (2) ATP and ADP perturbed the near UV CD only at spectral regions below 280 nm coinciding with the regions of their optical activity, (3) the optically inactive pyrophosphate caused no change in the near UV CD, and (4) myosin and HMM gave exactly the same results. These results suggest that myosin‐substrate interaction in vitro does not result in a delocalized conformational change.
Hormone receptor mobility and catecholamine binding in membranes. A theoretical model
Authors: McGuire Robert F. , Barber Roger 
Published: May 2005

Abstract[3 H]‐Catecholamine binding to intact cells, isolated cell membranes, and to several isolated macromolecules has been shown by several laboratories to be neither stereospecific nor inhibited by known β‐antagonists. Since additional evidence indicates that this binding is not an artifact (i.e. due neither to the binding of a catecholamine oxidation product nor hormone binding to a catabolic enzyme such as COMT), the question remains as to whether this represents binding to a bona fide membrane receptor. Because all ligands which bind strongly or compete for this binding possess a catechol group, one possible explanation is that the binding affinity is primarily determined by the catechol moiety, whereas the correct stereoisomer of the side chain is necessary to activate the receptor. Thus, although binding is a necessary condition for hormone action, the necessary and sufficient condition for activation of adenyl cyclase is both the catechol group and the correct stereoisomer of the side chain.A theoretical model is developed here to provide a quantitative basis for this hypothesis. This model extends the current concept of distinct subunits in the adenyl cyclase system by separating the receptors from the catalytic sites and placing them at separate locations within the membrane. Utilizing the spare receptor model of Furchgott, and the mobility of macromolecules within a “lipid sea,” the appropriate equations to predict both hormone binding and enzyme activation are derived. Using the observed affinity constants from catecholamine binding studies, it is then shown that this model can predict the experimental observations and hence explain the apparent dichotomy arising from binding and enzyme activation studies.
Perturbation of the chemotactic tumbling of bacteria
Authors: Taylor Barry L. , Koshland D. E. 
Published: May 2005

AbstractThe bacterial sensing system has been studied on three levels. First, a quantitative method has been devised for measuring the “action spectrum” of the bacterium in response to a sudden addition of attractant. Second, a technique has been developed for the rapid isolation of mutants defective in the transmission part of the sensing system. Third, a study of the effects of light on the transmission system reveals two components, one which generates tumbling and another which inhibits it.
Transport of amino acids in intact 3T3 and SV3T3 cells. Binding activity for leucine in membrane preparations of ehrlich ascites tumor cells
Authors: Cecchini Gary , Lee Mary , Oxender Dale L. 
Published: May 2005

AbstractTransport of amino acids into 3T3 and SV3T3 (SV40 virus‐transformed 3T3) cells was measured on glass cover slips. The 3T3 and SV3T3 cells contain both A (alanine preferring) and L (leucine preferring) systems for neutral amino acid transport. Initial rates of uptake of amino acids are about twofold higher in SV3T3 than in 3T3 cells. Other parameters measured, however, do not indicate marked differences in the transport of amino acids by the two cell types. L‐system amino acids, such as leucine, are subject to trans‐stimulation in both cell lines, whereas A‐system amino acids, such as alanine and glycine, are not. Leucine was transported to higher levels in confluent cells than in nonconfluent cells. Glycine, however, shows distinctly less transport activity as the cells become confluent.Ehrlich ascites cell plasma membranes were prepared and assayed for amino acid‐binding activity. Leucine‐binding activity was detected by equilibrium dialysis in Triton X‐100‐treated membrane preparations.
Conjugated polyene fatty acids as fluorescent membrane probes: Model system studies
Authors: Sklar Larry A. , Hudson Bruce S. , Simoni Robert D. 
Published: May 2005

AbstractThe use of conjugated polyene fatty acids as probes of membrane structure is examined. α‐ and β‐parinaric acid (cis, trans, trans, cis‐ and all trans‐9,11,13,15‐octadecatetraenoic acid) and synthetic lecithins containing an α‐parinaric acid chai in position 2 have been prepared, and their absorption and fluorescence properties have been determined. Their absorption spectra are at sufficiently long wavelength to be unobscured by cellular chromophores such as nucleotides and aromatic amin acids. Parinaric acid absorption does, however, overlap tryptophan emission which allows fluorescence energy transfer.Potential uses of these fluorescent probes are presented with studies on mode systems with known physical properties. Dipalmitoyl phosphatidylcholine exhibits a sharp phase transition 1° wide at 42° C, as monitored by the fluorescence intensit, of parinaric acid. The magnitude of the transition is independent of probe concentration, but the width of the transition and hysteresis are dependent upon such factors as the probe concentration and whether or not sonication is used in sample preparation. Using both fluorescence and absorption properties of the probe, we show that the addition of cholesterol to the dispersion broadens and decreases the magnitude of the transition. These results are interpreted in terms of a change in the polarizability of the acyl chains of a lipid bilayer as the bilayer undergoes a thermal transition.Lipid‐protein interactions are studied by the binding of α‐parinaric acid to bovine serum albumin. Fluorescence enhancement, absorption spectral shifts, and quenching of tryptophan fluorescence are observed when α‐parinaric acid binds to bovine serum albumin. Calculations based on these measurements are consistent with two binding sites of KB ∼ 108 (M−1) and three to four binding sites of KB ∼ 106 – 107 (M−1), similar to known values for the binding of other long‐chain fatty acids.Biosynthetic incorporation of β‐parinaric acid into the E. coli fatty acid auxotroph 30E βox− has been accomplished and phase transitions in cells and isolated phospholipids are shown.
The action of cytochalasin A on the in vitro polymerization of brain tubulin and muscle G‐actin
Authors: Himes Richard H. , Houston L. L. 
Published: May 2005

AbstractThe presence of cytochalasin A inhibits the self‐assembly of beef brain tubulin and rabbit muscle G‐actin in vitro and also decreases the colchicine binding of tubulin. Prior reaction of cytochalasin A with 2‐mercaptoethanol destroys its inhibitory effects. It is shown that cytochalasin A exerts its actions by reacting with sulfhydryl groups, possibly causing irreversible structural changes in the proteins. Cytochalasin B does not affect the tubulin assembly reaction.
Sarcomas routinely produced from putatively nontumorigenic Balb/3T3 and C3H/10T1/2 cells by subcutaneous inoculation attached to plastic platelets
Authors: Boone Charles W. , Jacobs Jerome B. 
Published: May 2005

AbstractThe Balb/3T3 and C3H/10T1/2 lines, noted for their marked postconfluence inhibition of proliferation and anchorage dependence, and frequently studied as nontumorigenic lines that are compared with tumorigenic sublines transformed with various agents, produced tumors within two to four months at low‐cell dosage (3 × 104 cells) when implanted subcutaneously attached to 1 × 5 × 10 mm polycarbonate platelets. Platelets alone did not produce tumors. The cultured Balb/3T3 tumor cells showed loss of both postconfluence inhibition of proliferation and anchorage dependence. Tumors arising form attached Balb/3T3 cells in (BALB/c × C57B1/6)F1 hybrids were shown to be transplantable to BALB/c but not to C57B1/6 mice, proving that the tumors were derived form Balb/3T3 and not from host cells. The tumors exhibited unique transplantation rejection antigens that did not cross‐react with each other. Scanning electronmicroscopy of Balb/3T3 cells and derive tumor cells on TeflonTeflon: Registered trademark of DuPont Plastics.
substrates (on which only the tumor cells and not the parent Balb/3T3 cells could grow) revealed that the two cell types were remarkably similar in appearance, except that the tumor cells were larger and showed many more microvilli that tended to concentrate over the nucleus. We conclude that Balb/3T3 cells and C3H/10T1/2 cells are preneoplastic and give rise to spontaneously transformed clones when implanted in vivo attached to a solid substrate.
Molecular composition and origin of substrate‐attached material from normal and virus‐transformed cells
Authors: Culp Lloyd A. 
Published: May 2005

AbstractThe proteins and polysaccharides which are left adherent to the tissue culture substrate after EGTA‐mediated removal of normal, virus‐transformed, and revertant mouse cells (so‐called SAM, or substrate‐attached material), and which have been implicated in the cell‐substrate adhesion process, have been characterized by SDS‐PAGE and other types of analyses under various conditions of cell growth and attachment. The following components have been identified in SAM: 3 size classes of hyaluronate proteoglycans; glycoprotein Co (the LETS glycoprotein); protein Ca (a myosin‐like protein); protein Cb (MW 85,000); protein C1 (MW 56,000, which is apparently not tubulin); protein C2 (actin); proteins C3–C5 (histones) which are artifactually bound to the substrate as a result of EGTA‐mediated leaching from the cell; and proteins Cc, Cd, Ce, and Cf. The LETS glycoprotein (Co) and Cd appear in newly‐synthesized SAM (which is probably enriched in “footpad” material – “footpads” being focal areas of subsurface membranous contact with the substrate) in greater relative quantities than in the SAM accumulated over a long period of time (which is probably enriched in “footprint” material – remnants of footpads left behind as cells move across the substrate). Co and Cd turn over very rapidly following short radiolabeling periods during chase analysis. The SAM's deposited during a wide variety of cellular attachment and growth conditions contained the same components in similar relative proportions. This may indicate well‐controlled and coordinate deposition of a cell “surface” complex involving the hyaluronate proteoglycans, the LETS glycoprotein, actin‐containing microfilaments with associated proteins, and a limited number of additional proteins in the substrate adhesion site. Evidence indicates that SAM is the remnant of “footpad” vesicles by which the cell adheres to the substrate and that EGTA treatment weakens the subsurface cytoskeleton, allowing these footpad vesicles to be pinched off from the rest of the cell. Three different models of cell‐substrate adhesion are presented and discussed.
Surface architecture of the plant cell: Biogenesis of the cell wall, with special emphasis on the role of the plasma membrane in cellulose biosynthesis
Authors: Montezinos David , Brown R. Malcolm 
Published: May 2005

AbstractCell wall structure and biogenesis in the unicellular green alga, Oocystis apiculata, is described. The wall consists of an outer amourphous primary layer and an inner secondary layer of highly organized cellulosic microfibrils. The primary wall is deposited immediately after cytokinesis. Golgi‐derived products contribute to this layer. Cortical microtubules underlie the plasma membrane immediately before and during primary wall formation. They function in maintaining the elliptical cell shape. Following primary wall synthesis, Golgi‐derived materials accumulate on the cell surface to form the periplasmic layer. This layer functions in the deposition of coating and cross‐linking substances which associate with cellulosic microfibrils of the incipient secondary wall. Secondary wall microfibrils are assembled in association with the plasma membrane. Freeze‐etch preparations of untreated, living cells reveal linear terminal complexes in association with growing cellulosic microfibrils. These complexes are embedded in the EF fracture face of the plasma membrane. The newly synthesized microfibril lies in a groove of the outer leaflet of the plasma membrane. The groove is decorated on the EF fracture face by perpendicular structures termed “ridges.” The ridges interlink with definitive rows of particles associated with the PF fracture face of the inner leaflet of the plasma membrane. These particles are termed “granule bands,” and they function in the orientation of the newly synthesized microfibrils. Microfibril development in relation to a coordinated multienzyme complex is discussed. The process of cell wall biogenesis in Oocystis is compared to that in higher plants.
Membrane protein alterations during the early stages of sporulation of bacillus subtilis
Authors: Goldman Robert C. 
Published: May 2005

AbstractMembrane protein alterations during the early stages of sporuloation were examined by polyacrylamide gel electrophoresis. Solubilized samples of the vegetative cell membrane (VCM), sporulation membrane fraction (SMF), and inner forespore membranes (IFM) were compared with respect to their protein compositions. The VCM contained 39 protein components, distinguishable as separate bands on gel electrophoresis, and these ranged in molecular weight from 16,000 to greater than 100,000. During the first 5 hr of sporulation, 6 of these 39 protein bands disappeared, 8 increased and 12 decreased in concentration, and 13 showed no discernible change. In addition, 15 new protein components were identified in the SMF during the fireist 5 hr. The new components consisted of 7 protein bands that were transiently associated with the SMF, and 8 proteins that persisted in the SMF from their time of appearance until at least T5 of sporulation. Comparison of the protein composition of the IFM with those of the VCM and SMF revealed that membrane protein alterations occur during sporulation.The turnover of H3‐tryptophan‐labeleld membrane protein was followed during growth and sporulation. During the 30 min of growth following a simple chase with excess unlabeled tryptophan, membrane protein appeared stable, whereas 5–10% of the nonmembrane protein turned over to acid‐soluble material. However, manipulation of the cells by dilution ito fresh medium, or centrifugation, as part of the chase procedure, resulted in elution of membrane protein to the cytoplasm. In contrast, proteins labeled during vegetative growth were always eluted to the cytoplasm during the first 2 hr of sporulation, and this was followed by a period of reassociation with the membrane fraction. The results are discussed with respect to membrane differentiation as it relates to spore development.
Ricinus communis toxin‐mediated inhibition of protein synthesis in cell‐free extracts of a toxin‐resistant variant mouse lymphoma cell line
Authors: Robbins James C. , Hunter Tony R. , Nicolson Garth L. 
Published: May 2005

AbstractRicinus communis agglutinin II (RCAII, ricin, toxin) at low concentrations inhibits protein synthesis in cell‐free extracts, but not in intact cells, of an RCAII‐resistant mouse lymphoma variant cell line. The concentration dependence of the inhibition by RCAII was the same in cell‐free extracts of both RCAII‐resistant variant and RCAII‐sensitive parental cells, while intact parental cells are 250 times more sensitive to RCAII toxicity. The onset of RCAII inhibition of cell‐free protein synthesis was extremely rapid in both cases, being complete in a few minutes. Under these condidtions RCAII inhibits protein synthesis in intact RCAII‐sensitive parental cells, but maximal inhibition requires several hours to occur. These results support our previous electron microscopic observations that the variant cells are defective in the uptake of RCAII by endocytosis at low toxin concentrations.
Two general classes of cytoplasmic actin filaments in tissue culture cells: The role of tropomyosin
Authors: Lazarides Elias 
Published: May 2005

AbstractIn the assembly of actin filaments that takes place during the spreading of a polulation of human lung cells, after trypsin detachment off the substratum and replating, tropomyosin exhibits a considrable lag in its association with the newly forming filament bundles; it begins to associate with them during the later stages of cell spreading as the actin filament bundles normally seen in interphase cells begin to organize. This lag is evident in a number of cell types that are spreading onto a substratum; it does not appear to be due to a selective degradation of this molecule during rounding up of the cells, since tropmyosin associates with the actin filament bundles after this lag even under conditions where the protein synthetic activity of the cell is inhibited to more than 95% by cycloheximide. The preferential binding of tropomyosin to fully assembled filament bundles but not to newly formed bundles of actin filaments suggests therefore the existence of two classes of action filaments: those that bind tropomyosin and those that do not. This selective localization of tropomyosin and those that do not. This selective localization of tropomyosin on actin filaments was further pursued by examining the localization of this molecule in membrane ruffles. The immunofluorescent results indicate that ruffling is an actin‐filament‐dependent, microtubule‐independent phenomenon. Tropomyosin is absent from membrane ruffles under a variety of circumstances where ruffling is expressed and, more generally, from any other cellular activity where actin filaments are expected to be in a dynamic state of reorganization or are required to be in a flexible configuraion. It is concluded that in tissue culture cells tropomyosin binds preferentially to actin filaments involved in structural support to confer rigidity upon them as well as aid them in maintaining a stretched phenotype. The absence of tropomyosin from certain motile phenomena where actin filaments are involved indicates that these classes of actin filaments are regulated by cytoplasmic mechanisms distinct from that by which tropomyosin (and troponin) mediates contractility in skeletal mulscle; it opens the possibility that different types of actin filaments enagaged in different cellular motile phenomenon in tissue culture cells may be regulated by a host of coexisting regulatory mechanisms, some as yet undetermined.
Isolation and purification of bacterial membrane proteins by the use of organic solvents: The lactose permease and the carbodiimide‐reactive protein of the adenosinetriphosphatase complex of escherichia coli
Authors: Altendorf Karlheinz , Lukas Margot , Kohl Brigitte , Müuller Clemens R. , Sandermann Heinrich 
Published: May 2005

AbstractTechniques for the solubilization and fractionation of integral membrane proteins have been developed in recent years. A small portion of membrane protein (about 2%, proteolipid fraction) will partition into chloroform or 1‐butanol, and, in several cases, these proteins retain functional activity. A virtually complete solubilization can be achieved at neutral pH by use of aprotic solvents, like hexamethylphosphoric triamide or N‐methylpyrrolidone.At relatively low concentrations (< 3 M) aprotic solvents inhibited β‐D‐galactoside transport by whole cells and the derivative membrane vesicles of Escherichia coli, but this inhibition could be largely reversed by a simple washing procedure. At higher concentrations of aprotic solvent (5–6 M), 50–80% of the total protein of lactose transport‐positive membrane vesicles was solubilized. When these extracts were added to intact lactose transport‐negative membrane vesicles, lactose transport was reconstituted, the required energy being provided by either respiration (e.g., addition of D‐lactate) or by a K+ diffusion potential established with the aid of valinomycin.The dicyclohexylcarbodiimide (DCCD)‐reactive subunit of the E. coli ATPase complex was found to partition into chloroform, and to be amenable to further purification in organic solvent. Ether precipitation and chromatography on DEAE‐cellulose and hydroxypropyl‐Sephadex G‐50 yielded an homogeneous polypeptide of an apparent molecular weight of 9,000.The purified and unlabeled DCCD‐reactive protein was incorporated into K+‐loaded liposomes, and a membrane potential was generated by the addition of valinomycin. There are indications that the DCCD‐reactive protein alone made the membrane specifically permeable for protons.