bluuebunny/crossref_metadata_2025_split huggingface dataset:
The differentiation of rat liver endoplasmic reticulum membranes: Apo–cytochrome P450 as a membrane protein
Authors: Siekevitz Philip 
Published: May 2005

AbstractThe proteins of washed microsomal membranes from adult rat liver were solubilized by 2% SDS and electrophoresed on polyacrylamide gels. Confirming earlier reports, a large Coomassie‐Blue staining band in the ∼50,000 MW region was identified as cytochrome P450 by four criteria: similar electrophoretic mobility to a purified cytochrome P450 preparation, an increase in this band after in vivo phenobarbital administration, a decrease in this band after in vivo allylisopropylacetamide administration, and direct specific binding of added purified heme to this region of a washed, unfixed gel. Although cyt P450 is not spectrally evident until just at the time of birth of the rats, a large band in this region was detectable in gels of microsomal membrane protein at all times, from three days before birth onward; this band also bound added heme after membrane proteins from fetal rat liver microsomes were electrophoresed on the gels. The conclusion was that apo‐cyt P450 is present in microsomal membranes at these times during differentiation, and that, regarding this protein, during differentiation heme is bound to the apo‐protein already there, concomitant with a synthesis of more cyt P450 molecules. The process of differentiation of this membrane type is also discussed.
Multiple forms of porcine pancreatic phospholipase A2: Isolation and specificity
Authors: Tsao Francis H. C. , Cohen Herschel , Snyder William R. , Kézdy Ferenc J. , Law John H. 
Published: May 2005

AbstractPorcine pancreatic phospholipase A2 was purified from commercial pancreatin by a method involving heat denaturation, trichloroacetic acid precipitation, and DEAE‐cellulose chromatography. Assaying the eluate of the chromatography step by a new titrimetric method using vegetable lecithin‐albumin emulsion as the substrate, several species of phospholipase A were found. Some of these went undetected when the conventional egg yolk emulsion assay was used. Two phospholipases A2 were isolated in a homogeneous form and shown to have similar chemical and physical properties. Catalytic specificity of the two enzymes differs remarkably toward lecithins in different emulsified states.
Genetic control of ribosome assembly
Authors: Sypherd Paul S. , Bryant Robert , Dimmitt Kathleen , Fujisawa Toshitaka 
Published: May 2005

AbstractThe assembly of E. coli ribosomes has been studied through the analysis of a new group of ribosome maturation mutants. These mutants, all blocked in a late stage in the maturation of 50S ribosomes, map at four different sites on the chromosome. These sites are distant from the known ribosomal protein sites at the str‐aro E region of the chromosome. The ribosome precursor particles of the mutants contain precursor‐type 23S RNA (p23 RNA) and 5S RNA. 43S particles of one of the mutants contain all but one of the normal complement of proteins. Precursor 43S particles from this mutant can be converted to particles with sedimentation values around 50S by incubation with extracts from either the wild‐type organism or from other mutants. This in vitro conversion process differs considerably from the process of ribosome reconstitution and indicates a role for extrinsic factors in the maturation of E. coli ribosomes.
The regulation of flagellar formation and function
Authors: Hilmen M. , Silverman M. , Simon M. 
Published: May 2005

AbstractThe basal structure of the flagellum controls both activity and assembly. In order to define the steps involved in these processes, genetic analysis was performed. Twenty genes were found to be required for the complete assembly and function of the organelle. FlaE controls the length of the hook, flaA is required both to maintain flagellar structure and for chemotaxis, and flaI plays a role in regulating the synthesis of the entire structure. Mutations mapping close to flal (the cfs mutations) release flagellar synthesis from control by catabolite repression.The basal structure was purified and isolated. On SDS acrylamide gel electrophoresis, it contained at least six distinguishable components. One major band corresponded to the hook subunit with an apparent molecular weight of 42,000 daltons. The others had apparent molecular weights of 60,000, 40,000, 28,000, 25,000, and 18,000 daltons. The genes that correspond to these polypeptides have not been identified.In exploring the role of the mot and che genes, assays were developed for the function of individual flagellar filaments. The filaments were found to rotate and rotation could be modulated by changing their direction. Chemotaxis results from the modulation of flagellar rotation. Using the rotation assay the response of nonmotile cells to attractants and repellents was followed.
Microtubule assembly: Some possible regulatory mechanisms
Authors: Olmsted J. B. , Marcum J. M. , Johnson K. A. , Allen C. , Borisy G. G. 
Published: May 2005

AbstractMicrotubule polymerization in vitro was examined using material purified from porcine brain tissue by a reversible temperature dependent assembly procedure, and was characterized by electron microscopy, viscometry, and sedimentation. The reaction was endothermic, colchicine sensitive, and occurred at neutral pH and moderate ionic strength. Divalent cations (calcium, magnesium) were inhibitory at millimolar concentrations, but stimulated polymerization at the micromolar level. Nucleoside triphosphates were required for assembly of purified subunits. As determined by quantitative sedimentation analyses, the reaction was an equilibrium process. Below a critical concentration of tubulin no assembly occurred. Analytical ultracentrifugation studies indicated that tubulin species with sO20, w of 6S and 30S were in equilibrium with each other, and that both were incorporated into microtubules. Electron microscopic analyses suggested that disc (or ring) structures might be intermediates in assembly, and that they were primarily utilized early in the polymerization process. Assembly could be seeded by mixing microtubular fragments from brain or flagella with brain microtubule subunits; depending on conditions of temperature and protein concentration, addition of subunits occurred either with unipolar or biased polar directionality. The possible significance of these properties of the polymerization reaction for control of assembly is discussed.
A molecular model of membrane excitability
Authors: Baumann Gilbert , Mueller Paul 
Published: May 2005

AbstractAlamethicin, monazomycin, or EIM induce electrical excitability in lipid bilayers. The voltage‐dependent gating displays all the characteristics observed in excitable cells and its basic features can be quantitatively described by the Hodgkin‐Huxley equations.A common molecular mechanism of membrane excitation has been postulated. It assumes that in the absence of an electrical field the channel‐forming molecules lie at the surface of the membrane. An applied potential tilts them from the surface into the hydrocarbon region of the bilayer. Once in this position the molecules diffuse laterally and form aggregates which act as channels for the flow of ions.In the case of alamethicin we assume that the molecule forms an elongated ellipsoid with two glutamic residues at one end, and a metal ion in four‐ or five‐fold coordination with peptide carbonyl oxygens at the other. An applied field pulls the cationic end through the membrane to the other side, while the glutamic residues hold the other end attached to the original surface. The molecules now span the membrane and aggregate, forming oligomeric channels in which most of the peptide carbonyls face toward the center, and the methyl groups outward.Monomers and dimers do not conduct and an individual channel can have different conductance values depending on the number of monomers in the aggregate and the resulting channel diameter. A quantitative description of this process matches observed gating kinetics, gating currents, and the single channel conductance increments. Without additional assumptions, inactivation follows directly from the aggregation process because with proper rate constants, the average degree of polymerization and therefore number of open channels goes through a maximum in time.The model may also apply to the excitation process of higher cells.
Calcium efflux from internally dialyzed squid axons: The influence of external and internal cations
Authors: Blaustein M. P. , Russell J. M. , De Weer P. 
Published: May 2005

AbstractInternal dialysis techniques have been used to examine the influence of external and internal cations on Ca efflux from ATP‐depleted squid axons. The main observation is that Ca efflux is promoted by external Na and inhibited by internal Na. The Na0 ‐dependent Ca efflux appears to be a function of [Na], and is also affected by the membrane potential; a 25 mV depolarization may cause as much as an e‐fold decrease in Ca efflux. These data are consistent with a counter‐transport exchange of 3Na+–for–1Ca2+. A Ca0‐dependent Ca efflux has also been observed; it is prominent in Na sea water or Le sea water, and is markedly diminished in choline sea water. This flux is consistent with the idea of a Ca‐Ca exchange diffusion process. Taken together, the Na0 ‐ and the Ca0 ‐dependent Ca effluxes fit a two‐site model for carrier‐mediated Ca transport; one site binds two Na+ or one Ca2+, while the second site can bind either one Na+ or one Li+. The data reported here suggest that both sites must be filled on the inward journey, but that only the Ca‐binding site need be occupied on the outward journey of the carrier. A mechanism of this type could derive sufficient energy from the Na and voltage gradients to maintain a [Ca2+]0/[Ca2+]i concentration ratio of about 104 in the absence of ATP. The present experiments do not, however, rule out the possible participation of a metabolically driven Ca transport mechanism in vivo.
Physical properties of the lipid phase of membranes from cultured animal cells
Authors: Wisnieski Bernadine J. , Huang Yoshie O. , Fox C. Fred 
Published: May 2005

AbstractESR analysis of membranes from cultured animal cells reveals a more complex lipid phase behavior than that displayed by ideal binary lipid systems. When endoplasmic reticulum membranes from LM cells are spin labeled with a nitroxide derivative of decane, 5N10, and scanned by ESR at 1° C‐intervals, the partitioning of 5N10 between the hydrocarbon and aqueous portions of the membrane suspension undergoes thermotropic changes at characteristic temperatures of 9°, 16°, 22°, 32°, and 38° C. Lipids extracted from these same membranes, however, exhibit only two characteristic temperatures, 16° and 35° C, and in this respect resemble binary lipid systems. The phase behavior of lipids in animal cell membranes is suggestive of an organized distribution of lipid which is disrupted by extraction. In support of this, mathematical treatment of the partitioning data indicates that four of these characteristic temperatures can define the boundaries (i.e., the t1 and th ) of two independent phase transitions in endoplasmic reticulum membranes. These results are similar to those of a physical treatment of data from plasma membranes of both mouse and chick cells in which the two monolayers appear to exist as independent physical entities with different physical properties. The most probable phase boundaries for the two monolayers of the endoplasmic reticulum membranes studied here are 16° and 32° C for one monolayer and 22° and 38° C for the other.
The effect of membrane‐lipid phase transitions on membrane structure and on the growth of acholeplasma laidlawii B
Authors: McElhaney Ronald N. 
Published: May 2005

AbstractThe physical state of the membrane lipids, as determined by fatty acid composition and environmental temperature, has a marked effect both on the temperature range within which A. laidlawii can grow and on the temperature coefficient of growth within the permissible temperature range. The minimum growth temperature under certain conditions is clearly defined by the lower boundary of the gel–to–liquid‐crystalline phase transition of the membrane lipids. The physical state of the membrane lipids can also influence the optimum and maximum growth temperatures. An a brupt increase in the temperature coefficient of growth is noted at temperatures between the phase transition boundaries. Both the absolute rates and the temperature coefficients of cell growth are similar for cells whose membrane lipids exist entirely or predominantly in the liquid‐crystalline state, but absolute growth rates decline rapidly and temperature coefficients increase when most of the membrane lipids become solidified. Some cell growth, however, can continue at temperatures at which less than 10% of the total lipid remains in the fluid state. Conversion of the membrane lipid from the liquid‐crystalline to the gel state is accompanied by a progressive aggregation of intramembranous protein particles. An appreciable heterogeneity in the physical state of the membrane lipids can apparently be tolerated by this organism without a detectable loss of membrane function.
Effect of membrane lipid composition and microtubule structure on lectin interactions of mouse lm cells
Authors: Rittenhouse Harry G. , Williams Robert E. , Fox C. Fred 
Published: May 2005

AbstractConcanavalin A (Con A) binding and Con A‐mediated hemadsorption to LM cells were found to decrease significantly at both 5–7°C and 15–19°C. The higher of these critical temperatures responds to a change in state of the membrane lipids and can be increased or decreased in cells where the membrane phospholipids contain less or more double bonds, respectively. The lower critical temperature for Con A binding or Con A‐mediated hemadsorption does not respond to these changes in membrane lipid composition. Though the amount of Con A bound to the cell surface is a determinant of Con A‐mediated agglutinability, the major components of the decreases in Con A‐mediated hemadsorption which occur at both these critical temperatures do not have their origin in the decreases in Con A binding which occur over these same temperature ranges – that is 5–7°C and 15–19°C.Con A‐mediated hemadsorption measured at 22°C was dramatically inhibited when LM cells were first incubated at 7°C or less. Reversal of this inhibition required 20–30 min of subsequent incubation at 22°C, indicating that factors other than membrane lipid “fluidity” are determinants of agglutinability. LM cells treated with the microtubule‐disrupting alkaloids colchicine, colcemid, or vinblastine at concentrations as low as 10−6 M were as much as fourfold more agglutinable with Con A. By contrast, lumicolchicine, an inactive derivative of colchicine, had a slight inhibitory effect on Con A‐mediated hemadsorption. Colchicine, vinblastine, or lumicolchicine treatment of LM cells did not alter the quantitative binding of labeled lectin. The results suggest that membrane lipid “fluidity” and the cell cytoskeleton (microtubule/microfilament system) are important determinants of lectin interactions with cell surfaces. The results are interpreted in terms of a model of cell–cell and cell–lectin interactions which assigns a central role to the Con A receptor.
Oxidation of lipids and membranes II: Effects of oxidants on cerebral white matter homogenates
Authors: Bubis J. J. , Wolman M. 
Published: May 2005

AbstractBovine cerebral white matter homogenates were oxidized by various oxidizing solutions of equal molarity and subsequently extracted with water. Most of the oxidants tested (K‐dichromate, FeCl3, H2O2, O2, and chloroperbenzoic, ascorbic, performic, and periodic acids) rendered the various myelin constituents less extractable than the constituents of unoxidized control homogenates. KMnO4, and to a lesser degree hemoglobin, rendered myelin constituents more extractable with water than those of the control homogenates. The findings indicate that most of the oxidants produced stabilization of the lamellar pattern, probably through cross‐linking and polymerization. KMnO4 and hemoglobin caused labilization and breakdown of the membranous structure. Proof that stabilization of membranes is caused by some oxidants and that fragmentation occurs by the action of KMnO4 and hemoglobin was obtained by the light‐scattering technique and by electron microscopy of the oxidized homogenates. Evidence obtained indicated that formation of hydrophobic end groups during oxidation favors polymerization, while prevalence of hydrophilic groups is associated with fragmentation.
Electron paramagnetic resonance and nanosecond fluorescence depolarization studies on creatine‐phosphokinase interaction with myosin and its fragments
Authors: Botts Jean , Stone Deborah B. , Wang Alice T. L. , Mendelson Robert A. 
Published: May 2005

AbstractRecent reports in the literature have indicated a physical association of creatinephosphokinase (CPK) with the tail portion of the myosin molecule. The present paper describes further studies on the interaction of CPK with myosin and myosin fragments, using the techniques of electron paramagnetic resonance (EPR) and nanosecond fluorescence depolarization. From EPR work, spin‐labeled CPK appears to interact with myosin, tail‐less myosin (heavy meromyosin [HMM]), and myosin heads (subfragment‐1 [S1]), the extent of interaction being proportional to the S1 content of myosin or its fragments. Spin‐labeled CPK did not evidence interaction with the headless myosin “rods”, with myosin tails (light meromyosin [LMM]), with S2 necks (which connect S1 to the rest of the myosin molecule), or with actin. When a fluorescent dye is directed to the essential ϵ‐amino group of CPK, nanosecond fluorescence depolarization studies indicate a substantial interaction with myosin, HMM, and S1, but very little with F‐actin. When the “fast‐reacting” thiol of the S1 moiety or the “essential thiol” of CPK was labeled with either a fluorescent dye or a spin label, no interaction between CPK and myosin (or S1) was detected.
Heat production and metabolism during the contraction of mammalian skeletal muscle
Authors: Kretzschmar K. M. 
Published: May 2005

AbstractMethods are described whereby initial processes of muscular contraction may be investigated in a mammalian preparation, the soleus muscle of the rat. Conditions are chosen so that recovery is avoided. An isometric tetanus is investigated and an energy balance sheet is drawn up. It is found that there is more heat evolved than can be accounted for in terms of measured chemical reaction. This discrepancy is discussed with reference to the similar results that have been obtained using frog muscle.
Energy conservation and uncoupling in mitochondria
Authors: Hatefi Youssef 
Published: May 2005

AbstractEnergy conservation and uncoupling in mitochondria are examined in the light of three important new findings: (a) Studies with the photoaffinity‐labeling uncoupler 2‐azido‐4‐nitrophenol have shown that mitochondria contain a specific uncoupler binding site (apparently a polypeptide of Mr = 30,000 ± 10%). (b) This site fractionates into an enzyme complex (complex V), which is capable of oligomycin‐ and uncoupler‐sensitive ATP‐Pi exchange. It is absent from electron transfer complexes I, III, and IV, which represent segments of the respiratory chain containing coupling sites 1, 2, and 3, respectively. (c) Trinitrophenol is a membrane‐impermeable uncoupler (uncouples submitochondrial particles, but not mitochondria) and a poor protonophore. There is an excellent correlation between the uncoupling potencies and the affinities of uncouplers for the mitochondrial uncoupler‐binding site. There is no correlation between uncoupling potency and protonophoric activity of uncouplers when a membrane‐permeable uncoupler is compared with a membrane‐impermeable one.
Exploring sites on mitochondrial ATPase for catalysis, regulation, and inhibition
Authors: Lardy Henry A. , Schuster Sheldon M. , Ebel Richard E. 
Published: May 2005

AbstractEvidence is presented that mitochondrial ATPase has two types of sites that bind adenine nucleotides. The catalytic site, C, binds the substrates ATP, GTP, or ITP and the inhibitor guanylyl imidodiphosphate (GMP‐PNP). A second type of site, R, binds ATP, ADP, adenylyl imidodiphosphate (AMP‐PNP), and the chromium complexes of ATP or ADP. All of these substances binding to the R site inhibit the hydrolysis of ATP in a competitive manner; their inhibition of hydrolysis of ITP and GTP is noncompetitive. GMP‐PNP inhibits oxidative phosphorylation in submitochondrial particles but AMP‐PNP does not. The localization on mitochondrial membranes of sites for the binding of various antibiotics that inhibit oxidative phosphorylation is discussed.
Studies of the oligomycin‐sensitive ATPase from yeast mitochondria. Reconstitution of ATP‐32Pi exchange in the presence of phospholipids
Authors: Ryrie Ivan J. 
Published: May 2005

AbstractA purified preparation of the oligomycin‐sensitive ATPase from yeast mitochondria has been shown to elicit an oligomycin‐ and uncoupler‐sensitive ATP‐32Pi exchange in the presence of phospholipids. Reconstitution was normally achieved by dialysis of an ATPase‐phospholipid‐cholate mixture. Following this procedure, vesicles with diameters between 200 and 1,500 Å were seen by electron microscopy. As in mitochondria, ATPase activity in the reconstituted system was stimulated by a range of uncouplers which inhibited ATP‐32Pi exchange. These and other findings suggest that the coupling mechanism may still be intact within the ATPase complex.
The effects of ADP on reverse electron flow and the oxygen exchange reactions catalyzed by bovine heart muscle submitochondrial particles
Authors: Mitchell Robert A. , Russo John A. , Lamos Candace M. 
Published: May 2005

Abstract1,N6‐Ethenoadenosine diphosphate (ϵ‐ADP) inhibits reverse electron flow (succinate → NAD+ driven by ATP) by competing with ATP, in contrast to ADP which we have shown previously to be a noncompetitive inhibitor. From these and other data it is concluded that the noncompetitive inhibition noted with ADP results from a combination of competitive inhibition plus non‐ or uncompetitive inhibition, the former occuring at a relatively nonspecific catalytic site and the latter at an extracatalytic site apparently quite specific for ADP.ADP, which stimulates ATP ⇌ H2O and Pi ⇌ H2O exchanges appears to be necessary for inhibition by arsenate of these exchanges. It is suggested that the ATP‐supported Pi ⇌ H2O exchange may be predominantly of the medium or intermediate type, depending on the concentrations of the Mg2+ complexes of ADP and Pi. Thus only exchanges involving medium ADP and Pi would be expected to show arsenate sensitivity.
Intermediate complex of ATP hydrolysis and synthesis by muscle proteins
Authors: Hotta K. 
Published: May 2005

AbstractMyosin catalyzed exchange between 32Pi and ATP in reaction medium during its enzymatic hydrolysis of ATP only by a very small amount. Addition of actin increased to a great extent the rate of incorporation of 32Pi in the presence of Mg. Glycerinated smooth muscle fibers also exhibited the ability to exchange 32Pi and ATP upon the application of external force (repeated stretching and releasing). A schematic mechanism of the action of actin and external force on acceleration of 32Pi incorporation is proposed and the importance of the M*‐ADP complex for force generation is suggested.