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1.
Peter R. Allegrini Guus van Scharrenburg Gerard H. De Haas Joachim Seelig 《生物化学与生物物理学报:生物膜》1983,731(3):448-455
1-Palmitoyllysophosphatidylcholine has been mixed in equimolar amounts with specifically deuterated palmitic acid and the structural properties of the lipid/water phase have been studied by 2H- and 31P-nuclear magnetic resonance. The order profile of the free palmitic acid is very similar to that of the parent compound at temperatures above the gel-to-liquid crystal phase transition. The bending of the chain which is typical for diacyl lipids is not observed for the free palmitic acid. The mixture of lysolipid and palmitic acid exhibits well-defined quadrupole splittings even at temperatures below the gel-to-liquid crystal phase transition. Hence it is possible for the first time to establish an order profile in the gel-state of the lipid bilayer phase. Between carbon atoms 5 to 12 the palmitic acid chain is found to assume the extended all-trans conformation with a very small contribution from gauche defects. Towards the methyl terminal a distinct increase in the gauche probability can be noted. The motion of the phosphocholine headgroup was also studied by 2H- and 31P-NMR using selectively deuterated 1-palmitoyllysophosphatidylcholine. The headgroup has a considerably larger motional freedom in the mixture of lysolipid and palmitic acid than in . In addition, the average headgroup conformations are also different in the two systems. 相似文献
2.
The action of ATP and its analogs as well as the effects of alkali ions were studied in their action on the ouabain receptor. One single ouabain receptor with a dissociation constant () of 13 nM was found in the presence of () and (). pH changes below pH 7.4 did not affect the ouabain receptor. Ouabain binding required Mg2+, where a curved line in the Scatchard plot appeared. The affinity of the receptor for ouabain was decreased by K+ and its congeners, by Na+ in the presence of (), and by ATP analogs (ADP-C-P, ATP-OCH3). Ca2+ antagonized the action of K+ on ouabain binding. It was concluded that the ouabain receptor exists in a low affinity (Rα) and a high affinity conformational state (Rβ). The equilibrium between both states is influenced by ligands of . With 3 mM Mg2+ a mixture between both conformational states is assumed to exist (curved line in the Scatchard plot). 相似文献
3.
The specific synthesis of F mRNA directed by the F gene carried on the specialized transducing bacteriophage F, performed , is described with the use of an S180 extract from a strain carrying R?. Synthesis of F mRNA is biphasic at approximately 7 minutes. The regulation of F mRNA synthesis by the specific arginine holorepressor present in an S180 extract prepared from a strain carrying the R+ allele is described. 相似文献
4.
G.E. Breitwieser 《生物化学与生物物理学报:生物膜》1982,689(3):457-463
(1) A membrane fraction enriched in (EC 3.6.1.3) was obtained from optic ganglia of the squid (Loligo pealei) by density gradient fractionation of membranes followed by treatment with either SDS or Brij-58. The resulting membrane had an specific activity of approx. 2 units/mg and was ouabain-sensitive. (2) The had a for ATP of and a pH optimum of 7.0. It was inhibited by ouabain with a of . (3) Optimum monovalent cation concentrations were: 240 mM NaCl, 60 mM KCl, tested with . (4) The Mg2+ dependence of hydrolysis varied with the absolute ATP concentration. At 3 mM ATP, the for Mg2+ was , and at 6 mM ATP, the was . High levels of Mg2+ caused inhibition of hydrolysis. (5) The interactions of Na+ and K+ were examined over a range of conditions. K+ levels caused modulations in the Na+ dependence in the range of 1–150 mM. (6) The prepared from squid optic ganglion displays properties similar to those of the sodium pump in injected nerves. 相似文献
5.
《Biochimica et Biophysica Acta (BBA)/General Subjects》1983,757(3):342-351
The existence of metabolically distinct pools of in Saccharomyces cerevisiae and isolated rat hepatocytes was investigated. Utilizing a relatively long labeling period with [methyl-14C]methionine, a metabolically ‘stable’ pool was labeled. A subsequent short labeling with [methyl-3H]methionine selectively labeled a putative metabolically ‘labile’ pool. The existence of these distinguishable pools was ascertained by following the 3H and 14C label disappearance in during the chase-period in label-free media containing cycloleycine to prevent futher synthesis of . In both yeast and hepatocytes, the ratio in decreased sharply. The individual 3H and 14C decrease in showed values of 3 and 8 min for yeast and 4 and 18 min for hepatocytes. The results strongly indicate that at least two metabolically distinct pools actually do exist in both systems. Subcellular fractionation revealed that the ‘labile’ pool exist in the cytosol for both yeast and hepatocytes while the ‘stable’ pool exists in the vacuolar and the mitochondrial fraction for the yeast and hepatocytes respectively. The pools were also studied in normal yeast under anaerobic chase condition and petite mutant yeast. Sharply contrasting with aerobically chased normal yeast, both showed closely parallel 3H and 14C decreases in . 相似文献
6.
Pretreatment of Chang liver cells with (0.5 or 1 mM) stimulated Na+-independent uptake of leucine at low concentrations (?1 mM). The stimulatory effect of on the uptake of leucine measured in Na+-replete medium was completely blocked by the addition of (5 mM), which shows that the L system participates in the stimulation. The Na+-dependent uptake of glycine was depressed by pretreatment. The stimulation of the Na+-independent component of leucine uptake continued for at least 30 min after treatment, while the inhibition of glycine uptake was progressive with time and the Na+-dependent uptake of leucine became depressed later, after the treatment. It has been demonstrated that treatment of cells with is capable of increasing the Na+-independent influx of leucine and at the same time slightly decreasing the efflux of it. These results suggest that attacks the Na+-independent system of amino acid transport at the reactive SH groups(s) of relevant protein(s) in favor of specific activation of that system in this cell. 相似文献
7.
Basolateral membranes isolated from hog kidney cortex, enriched 12- to 15-fold in (Na+ + K+)-ATPase activity, were 80% oriented inside-out as determined by assay of oubain-sensitive (Na+ + K+)-ATPase activity before and after opening of the membrane vesicle preparation with a mixture of deoxycholate and EDTA. In these membrane preparations 80% of total phosphatidylethanolamine was accessible to trinitrophenylation by trinitrobenzenesulfonic acid at 4°C, while at 37°C all of phosphatidylethanolamine fraction was chemically modified. Phospholipase C treatment resulted in hydrolysis of 80% phosphatidylethanolamine, 40% phosphatidylcholine and 35% of phosphatidylserine. Sphingomyelinase treatment resulted in 20% hydrolysis of sphingomyelin, presumably derived from right-side-out oriented vesicles. Results indicate that phosphatidylethanolamine is oriented exclusively on the outer leaflet of the lipid bilayer of inside-out oriented vesicles. Methylation of phospholipids in basolateral membranes with resulted in the three successive methylation of ethanolamine moiety of phosphatidylethanolamine to phosphatidylcholine. The for was 1·10?4 M with an optimum pH 9.0 for the formation of all three methyl derivatives. Mg2+ was without any effect between pH 5 and 10. Basolateral membranes incubated in the presence of methyl donor, , exhibited increased (12–15%) (Ca2+ + Mg2+)-ATPase activity and increased ATP-dependent uptake of calcium. ATP-dependent calcium uptake in these vesicles was insensitive to oligomycin and ouabain but was abolished completely by 50 μM vanadate. The increase in ATP-dependent calcium uptake was due to an increase in and not due to a change in for Ca2+. Preincubation of membranes with , a methyltransferase inhibitor, abolished the stimulatory effect of phospholipid methylation on calcium uptake. Phospholipid methylation at both low and high pH did not result in a change in bulk membrane fluidity as determined by the fluorescence polarization of diphenylhexatriene. These results suggest that phospholipid methylation may regulate transepithelial calcium flux in vivo. 相似文献
8.
Klaus Gietzen Peter Adamczyk-Engelmann Andreas Wüthrich Anka Konstantinova Hermann Bader 《生物化学与生物物理学报:生物膜》1983,736(1):109-118
Compound , a condensation product of N-methyl-p-methoxyphenethylamine with formaldehyde, is composed of a family of cationic amphiphiles differing in the degree of polymerization. Compound was found to be a potent inhibitor of the calmodulin-activated fraction of brain phosphodiesterase and red blood cell Ca2+-transport ATPase, with IC50 values of 0.3 and 0.85 μg/ml, respectively. However, the basal activity of both enzymes is not at all suppressed by the drug at concentrations up to 300 μg/ml. Inhibition of Ca2+ transport into inside-out red blood cell vesicles by compound follows a similar pattern in that basal, calmodulin-independent, transport is also not affected by the drug. Kinetic analysis revealed that the stimulation of Ca2+-transport ATPase induced by calmodulin is inhibited by compound according to a competitive mechanism. It was demonstrated that the inhibitory constituents of compound bind to calmodulin in a Ca2+-dependent fashion. Comparison of the specificity of several anti-calmodulin drugs showed that compound is the most specific inhibitor of the calmodulin-dependent fraction of red blood cell Ca2+-transport ATPase that has been described hitherto. In addition, compound was found to be a rather specific inhibitor of the calmodulin-induced activation of Ca2+-transport ATPase when compared with the stimulation induced by an anionic amphiphile or by limited proteolysis. Half-maximal inhibition of the activity stimulated by oleic acid or mild tryptic digestion required 8- and 32-times higher concentrations of compound , respectively, compared with the calmodulin-dependent fraction of the ATPase activity. Moreover, calmodulin-independent systems as rabbit skeletal muscle sarcoplasmic reticulum Ca2+-transport ATPase or calf cardiac sarcolemma (Na+ + K+)-transport ATPase are far less influenced by compound as compared with trifluoperazine and calmidazolium. Because of its high specificity compound is proposed to be a promising tool for studying calmodulin-dependent processes. 相似文献
9.
Mohammad Ali Ghalambor Christopher D. Warren Roger W. Jeanloz 《Biochemical and biophysical research communications》1974,56(2):407-414
Incubation of UDP-[14C]--acetylglucosamine with calf pancreas microsomes in the presence of Mn++ and potassium thiocyanate gave a labeled glycolipid, tentatively identified as 1-2-acetamido-2-deoxy--glucosyl 2-dolichyl pyrophosphate on the basis of cochromatography with synthetic 1-2-acetamido-2-deoxy-α--glucopyranosyl 2-dolichyl pyrophosphate, similar chemical and enzymic hydrolyses of the biosynthetic and synthetic compounds, and stimulation of the biosynthesis by addition to the incubation mixture o dolichyl phosphate or a crude lipid fraction extracted from microsomes. 相似文献
10.
The binding of the gamma labeled neuroleptic, 77Br--bromosprioperidol, in the rat brain was examined . This binding parallels the binding of 3H-spiroperidol, in that binding is especially high in dopaminergically innervated areas, is saturable, and is displaced by high doses of unlabeled spiroperidol (1–5). Thus, 77Br--bromospiroperidol is a suitable ligand for use in gamma ray imaging techniques for monitoring of receptor binding. 相似文献
11.
The block of the Na+ current by was studied in crayfish giant axons under axial wire voltage-clamp conditions. Standard kinetic analysis of the Na+ currents was undertaken to test the hypothesis that the block of the Na+ current could be accounted for on the basis of changes in the voltage dependence of the kinetic parameters. Alterations in the membrane dipolar potential arising from rearrangement of membrane lipids would be the anticipated source of changes in the voltage dependence. Although some changes in voltage dependence did evolve with the block by , the changes were not of sufficient magnitude to account for the block. In conclusion, although higher concentrations of produced shifts along the voltage axis of the kinetic parameters, direct blocking action of on the channel appears to be the most important mechanism of the block. 相似文献
12.
An ATPase is demonstrated in plasma membrane fractions of goldfish gills. This enzyme is stimulated by Cl? and HCO3?, inhibited by SCN?.Biochemical characterization shows that HCO3? stimulation () is specifically inhibited in a competitive fashion by SCN? (). The residual Mg2+-dependent activity is weakly is weakly affected by SCN?.In the microsomal fraction chloride stimulation of the enzyme occurs in the presence of HCO3? (); no stimulation is observed in the absence of HCO3?. Thiocyanate exhibits a mixed type of inhibition () towards the Cl? stimulation of the enzyme.Bicarbonate-dependent ATPase from the mitochondrial fraction is stimulated by Cl?, but this enzyme has a relatively weak affinity for this substrate (). 相似文献
13.
John A. Jacquez 《生物化学与生物物理学报:生物膜》1973,318(3):411-425
The Michaelis-Menten parameters, and of the initial 1-min fluxes of uptake of l-phenylalanine and of α-aminoisobutyric acid were determined for extracellular concentrations of Na+ ranging from 0.5 to 110 mequiv/l for Ehrlich ascites tumor cells. The maximal initial flux, , decreased with decrease in extracellular Na+ for both α-aminoisobutyric acid and phenylalanine but the for α-aminoisobutyric acid increased markedly as the Na+ concentration fell whereas the for phenylalanine decreased. Cycloleucine behaved like phenylalanine.The data provides strong evidence that the Na+-independent flux of phenylalanine is an exchange diffusion flux that can be varied by changing the intracellular level of amino acids such as phenylalanine. For phenylalanine, cyclolcucine, and methionine this exchange diffusion flux appears to be additive with the Na+-dependent initial flux. α-Aminoisobutyric acid also has an exchange diffusion that is Na+-independent but it has a high and is not additive with the Na+-dependent flux. 相似文献
14.
Quercetin inhibited a dog kidney ( preparation without affecting for ATP or for cation activators, attributable to the slowly-reversible nature of its inhibition. Dimethyl sulfoxide, a selector of E2 enzyme conformations, blocked this inhibition, while the K+-phosphatase activity was at least as sensitive to quercetin as the ( activity, all consistent with quercetin favoring E1 conformations of the enzyme. Oligomycin, a rapidly-reversible inhibitor, decreased the for ATP and the for cation activators, and its inhibition was also diminished by dimethyl sulfoxide. Although oligomycin did not inhibit the K+-phosphatase activity under standard assay conditions, a reaction presumably catalyzed by E2 conformations, its effects are nevertheless accommodated by a quantitative model for that reaction depicting oligomycin as favoring E1 conformations. The model also accounts quantitatively for effects of both dimethyl sulfoxide and oligomycin on , for substrate, and for K+, as well as for stimulation of phosphatase activity by both these reagents at low K+ but high Na+ concentrations. 相似文献
15.
This investigation was principally undertaken to test the ionic gradient hypothesis as applied to active uptake in the rabbit kidney cortical slice preparation. Efflux of from the slice was shown to be independent of external Na+ concentration. Transferring slices from a low sodium preincubation to a high sodium incubation medium containing increased intracellular concentrations of both Na+ and K+, and accumulation occurred. Transferring slices from a low sodium preincubation to a high sodium incubation medium containing ouabain and resulted in a net increase in intracellular Na+ concentration but no accumulation occurred. Different combinations of preincubation and incubation media gave a high to low array of intracellular Na+ concentrations and these directly reflected their respective uptake. These results suggest that the Na+-gradient hypothesis does not adequately explain the transport of organic acids in rabbit kidney. These results also suggest that Na+ possibly has an intracellular role through its stimulation of channeled to energizing the accumulative mechanism. 相似文献
16.
incorporates 3H or 14C from 3H- or 14C-labeled ethanolamine into an -alkali soluble, alcohol -insoluble fraction obtained from cell walls. Dansyl ethanolamine was isolated from this alcohol-insoluble fraction following dansylation and hydrolysis. The alcohol-insoluble material was non-dialyzable and contained galactofuranosyl, glucosyl, phosphoryl, amino acyl and variable quantities of uronosyl residues. The lack of detectable quantities of mannosyl residues in this material suggests that the galactofuranosyl-containing cell wall polymer is distinct from the peptidophosphogalactomannan which is obtained from culture filtrates of . (Gander ., (1974) J. Biol. Chem. , 2063). 相似文献
17.
The Na+-independent leucine transport system is resolved into two components by their different affinity ( about 44 μM and 8.0 mM) for leucine in the Chang liver cell. Treatment of the cells with (1 mM) specifically stimulates the high-affinity component of the Na+-independent system by greatly increasing its value, whereas the value of the low-affinity component is markedly lowered. The stimulatory effect of on leucine transport is reduced by prior treatment of the cells with 2,4-dinitrophenol, but this phenomenon seems to be irrelevant to the ATP-depleting action of the uncoupler. The treatment with 2,4-dinitrophenol has been found not to be inhibitory on the subsequent Na+-independent leucine uptake itself. Treatment with dibucaine, a phospholipid-interacting drug, also reduces to varying degrees (depending on its concentration) the stimulatory effect of on the subsequent leucine uptake, although pretreatment with dibucaine can stimulate the Na+-independent leucine uptake itself. We conclude that the stimulatory effect of on leucine transport is not correlated with the energy level of cell, but involves the perturbation of the membrane bilayer structures. 相似文献
18.
H. Engelberg-Kulka L. Dekel M. Israeli-Reches 《Biochemical and biophysical research communications》1981,98(4):1008-1015
The regulation of the synthesis of operon enzymes was studied in streptomycin-resistant mutants temperature-sensitive for UGA suppression by normal tRNATrp. Our mutants carry a allele that when transferred to a different genetic background causes repression of trp operon enzyme synthesis at both low (35°C) and high (42°C) temperatures; however, in our mutants with an excess of tryptophan and at increased temperatures enzyme synthesis is derepressed. Based on our results and the sequence data of the R gene [Singleton et al. (1980) Nucleic Acids Res., 8, 1551–1560], we offer a model for the involvement of the limited misreading of UGA codons by normal charged tRNATrp in the autogenous regulation of the R gene expression. The UGA readthrough process may be a regulatory amplifier of the effect of tryptophan starvation. 相似文献
19.
Michael D.P. Boyle 《Biochemical and biophysical research communications》1981,103(3):856-862
Zinc sulphate in the range of 10?4 to 2×10?5 M prevents the binding of to antigen antibody complexes, and the initation of the cascade of events in the classical complement pathway leading to cell lysis. Other heavy metals, Co++, Cd++, Cu++, or Mn++ were without effect in this concentration range. Zinc was ineffective when added after was bound and failed to displace which was already bound to antigen antibody complexes. The ability of zinc to regulate the binding of the zymogen or activated form of to antigen-antibody complexes represents a new method of controlling the initiation of the classical complement pathway. 相似文献
20.
The ATP/ADP exchange is shown to be a partial reaction of the by the absence of measurable nucleoside diphosphokinase activity and the insensitivity of the reaction to , -di(adenosine-5′) pentaphosphate, a myokinase inhibitor. The exchange demonstrates an absolute requirement for Mg2+ and is optimal at an ADP/ATP ratio of 2. The high ATP concentration () required for maximal exchange is interpreted as evidence for the involvement of a low affinity form of nucleotide site. The ATP/ADP exchange is regarded as evidence for an ADP-sensitive form of the phosphoenzyme. In native enzyme, pre-steady state kinetics show that the formation of the phosphoenzyme is partially sensitive to ADP while modification of the enzyme by pretreatment with 5,5′-dithiobis(2-nitrobenzoic acid) (DTNB) in the absence of Mg2+ results in a steady-state phosphoenzyme population, a component of which is ADP sensitive. The ATP/ADP exchange reaction can be either stimulated or inhibited by the presence of K+ as a function of pH and Mg2+. 相似文献