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1.
Sarcoplasmic reticulum membranes from skeletal muscle contain a proteolipid () which reduces both the nonspecific ion and water permeabilities of artificial planar phospholipid bilayers. The proteolipid does not show any ionophoric effect or specific pore formation for Ca2+. The a.c. capacitance of the bilayers is unaffected whereas the refractive index is increased by the presence of proteolipid. The results support the view that the proteolipid interacts with the phospholipids in the bilayer interior and causes a condensation in the packing of the alkyl chains. 相似文献
2.
The interactions between calmodulin, ATP and Ca2+ on the red cell Ca2+ pump have been studied in membranes stripped of native calmodulin or rebound with purified red cell calmodulin. Calmodulin stimulates the maximal rate of by 5–10-fold and the rate of Ca2+-dependent phosphorylation by at least 10-fold. In calmodulin-bound membranes ATP activates along a biphasic concentration curve (), but in stripped membranes the curve is essentially hyperbolic (). In calmodulin-bound membranes Ca2+ activates at low concentrations () in stripped membranes the apparent Ca2+ affinities are at least 10-fold lower.The results suggest that calmodulin (and perhaps ATP) affect a conformational equilibrium between E2 and E1 forms of the Ca2+ pump protein. 相似文献
3.
Temperature-jump-induced absorption changes of bromocresol purple in chloroplast suspensions in the dark were studied. After a rapid rise in temperature (<10 μs), a slow absorbance decrease of bromocresol purple () following a fast absorbance decrease of chloroplasts and bromocresol purple () was observed. The slow absorbance decrease corresponds to acidification of the suspending medium, indicating H+ efflux from chloroplasts after the temperature jump. Nigericin and gramicidin D suppressed the slow absorbance change completely in the presence of 10 mM KCl, while valinomycin did not affect it. The fast absorbance change was not affected by the above ionophores. 3-(3,4-dichlorophenyl)-1,1-dimethylurea also diminished the slow absorbance change. 相似文献
4.
Charge-pulse current-relaxation studies have been performed with lipid bilayer membranes in the presence of the hydrophobic ion dipicrylamine. From the analysis of the relaxation times and amplitudes the translocation rate constant of dipicrylamine as well as the partition coefficient β between membrane surface and water could be evaluated. In a first series of experiments membranes made from monoolein or dioleoylphosphatidylcholine in a number of different solvents were studied, as well as virtually solvent-free bilayer membranes made from monolayers. The thickness of the hydrocarbon layer of these membranes varied between 5.0 and 2.5 nm. While β was almost insensitive to variations in , a strong decrease of with increasing membrane thickness was found; the observed dependence of on approximately agreed with the theoretically expected influence of membrane thickness on the height of the dielectric barrier. No specific differences between Mueller-Rudin films and solvent-free (Montal-Mueller) membranes other than differences in thickness were found. In a further series of experiments the chemical structure of the lipid was systematically varied (number and position of double bonds in the hydrocarbon chain, nature of the polar head group). The translocation rate constant was much larger in phosphatidylethanolamine membranes than in phosphatidylcholine membranes. A strong increase of was found when the number of double bonds in the hydrocarbon chain was increased from one to three. These changes were discussed in terms of membrane fluidity and dielectric barrier height. Much higher values of were observed in lipids with ester linkage between hydrocarbon chain and glycerol backbone, as compared with the corresponding ether analogs. This finding is qualitatively consistent with determinations of dipolar potentials in monolayers of ester and ether lipids. When cholesterol is added to phosphatidylcholine membranes, the translocation rate constant increases up to five-fold, while the partition coefficient β remains virtually constant. The variation of in this case can be largely accounted for by a decrease in membrane thickness and a concomitant reduction in dielectric barrier height. In membranes made from the negatively charged lipid phosphatidylserine the partition coefficient of dipicrylamine strongly increased with ionic strength, as expected from the Gouy-Chapman theory of the surface potential. 相似文献
5.
The permeability of the lysosomal membrane to small anions and cations was studied at 37°C and pH 7.0 in a lysosomal-mitochondrial fraction isolated from the liver of untreated rats. The extent of osmotic lysis following ion influx was used as a measure of ion permeancy. In order to preserve electroneutrality, anion influx was coupled to an influx of K+ in the presence of valinomycin, and cation influx was coupled to an efflux of H+ using the protonophore . Lysosomal lysis was monitored by observing the loss of latency of two lysosomal hydrolases.The order of permeability of the lysosomal membrane to anions was found to be and that to cations . These orders are largely in agreement with the lyotropic series of anions and cations.The implications of these findings for the mechanism by means of which a low intralysosomal pH is produced and maintained are discussed. 相似文献
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7.
The kinetic α-secondary deuterium isotope effect, , for the pH-independent hydrolysis of nicotinamide riboside, yielding nicotinamide and ribose, in water at 25 ° is 1.14, establishing that this reaction proceeds with unimolecular substrate decomposition to yield a carboxonium ion, or related species, in the rate-determining step. Surprisingly, the corresponding isotope effect for the base-catalyzed decomposition of the same substrate is 1.12, a value indicating considerable sp2 character at the Cl′ position in the transition state for this reaction. A similar result, , was obtained for base-catalyzed hydrolysis of NAD+. The kinetic alpha deuterium isotope effect for the pig brain NAD glycohydrolasecatalyzed hydrolysis of nicotinamide riboside is 1.08. This value suggests that CN bond cleavage to form an intermediate carboxonium ion, or structurally related species, is at least partially rate-determining. In contrast, the corresponding value for the hydrolysis of this substrate catalyzed by Escherichia coli nicotinamide ribonucleotide glycohydrolase is very near unity, a result consistent with several interpretations including a rate-determining enzyme isomerization reaction. 相似文献
8.
M J Modak S L Marcus L F Cavalieri 《Biochemical and biophysical research communications》1973,55(1):1-7
Incubation of liver microsomes with cytochrome , purified after solubilization with detergents, caused an effective incorporation of the cytochrome into the microsomal membranes. The incorporated cytochrome was reducible by NADH and could not be removed by repeated washing with 0.3 M KCl or 10 mM EDTA. The incorporation was much more efficient at 37°C than at 0°C. Trypsin-solubilized cytochrome , which lacks the hydrophobic tail of the native protein, could not be inserted into the membranes. These findings confirm the view that the hydrophobic tail of the cytochrome molecule is responsible for its tight binding to the microsomal membranes. 相似文献
9.
A method is described to measure the oxygen diffusion-concentration product, , at any locus that can be probed or labeled using nitroxide radicals. The method is based on the dependence of the spin-lattice relaxation time of the spin label on the bimolecular collision rate with oxygen. Strong Heisenberg exchange between spin label and oxygen contributes directly to of the spin label, while dipolar interactions are negligible. Both time-domain and continuous wave saturation methods for studying are considered. The method has been applied to phospholipid liposomes using fatty acid spin labels. A discontinuity in at the main phase transition was observed. 相似文献
10.
This paper presents an interpretation of fluorescence polarization measurements in lipid membranes which are labelled with the apolar probe 1,6-diphenyl-1,3,5-hexatriene. The steady-state fluorescence anisotropy, , is resolved into a fast decaying or kinetic component, , and an infinitely slow decaying or static component, . The latter contribution, which predominates in biological membranes, is exclusively determined by the degree of molecular packing (order) in the apolar regions of the membrane; is proportional to the square of the lipid order parameter. An empirical relation between and is presented, which is in agreement with a prediction based on a theory of rotational dynamics in liquid crystals. This relation enabled us to estimate a lipid structural order parameter directly from simple steady-state fluorescence polarization measurements in a variety of isolated biological membranes. It is shown that major factors determining the order parameter in biomembranes are the temperature, the cholesterol and sphingomyelin content and (in a few systems) the membrane intrinsic proteins. 相似文献
11.
R.P. Brivio-Haugland S.L. Louis K. Musch N. Waldeck M.A. Williams 《生物化学与生物物理学报:生物膜》1976,433(1):150-163
To determine whether changes in unsaturation of fatty acids in rat liver plasma membranes might alter activities of membrane-associated enzymes, liver plasma membranes were prepared from rats fed purified diets lacking or supplemented with essential fatty acids. Two methods of membrane purification were used. A similar degree of purification was obtained with both methods for both depleted and control membranes, as indicated by marker enzyme purification. The proportion of essential fatty acids of the linoleate series was significantly lower in phospholipids from depleted rats. The specific activity of 5′-nucleotidase was lower, and the activity, and apparent for total were higher in the depleted liver plasma membranes. Arrhenius plots of total ATPase activity showed a discontinuity at the same temperature for both the depleted and control membranes. Activity with the depleted membranes was higher at all temperatures tested. Supplementation of deficient rats with a source of essential fatty acids (corn oil) restored and apparent values to normal. Adenylate cyclase activity in the presence of fluoride, glucagon or glucagon plus GTP was significantly lower in the depleted plasma membranes. 相似文献
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13.
Charge-pulse relaxation experiments with the negatively charged lipophilic ions, dipicrylamine and tetraphenylborate, (as well as with the positively charged carrier system Rb+-valinomycin) have been carried out in order to study the influence of sterols on the ion transport through the lipid bilayer membrane. The mol fraction of the sterols (cholesterol, epicholesterol, ergosterol, stigmasterol, dihydrocholsterol, epicoprostanol and cholesterololeate) as referred to total lipid was varied in a wide range (mol fractions 0–0.8).The monoolein/sterol or dioleoylphosphatidylcholine/sterol mixtures were dissolved in -hexadecane in order to minimize effects of the sterol on the membrane thickness.Cholesterol had a strong influence on the transport of the lipophilic ions. Its incorporation into monoolein membranes increased the rate constant of translocation up to 8-fold, but incorporation into phosphatidylcholine membranes had virtually no influence on . The other sterols with one hydroxy group and cholesterololeate had no influence on the rate constant or the partition coefficient β. The results are discussed on the basis of a possible change of dipole potential of the membrane caused by cholesterol and its derivatives.In the case of valinomycin-mediated Rb+ transport only cholesterol had a strong influence on transport properties. The rate constants of association () as well as the rate constants of translocation of the complex () and of the free carrier () were reduced by incorporation of cholesterol up to eight-fold. The decrease of and are possibly caused by a decrease of membrane fluidity, whereas the decrease of may be due to an increase of surface potential. The different action of cholesterol on the two transport systems is discussed under the assumption that the adsorption plane of the lipophilic ion is located more towards the aqueous side and that of the ion-carrier complexes more towards the hydrocarbon side of the dipole layer. 相似文献
14.
N-(2-dimethylaminoethyl)acetohydroxamic acid was synthesized. This compound, which incorporates a dimethylamino group as a second functionality into the hydroxamic acid molecule, catalyzes the hydrolysis of p-nitrophenyl acetate faster than acetohydroxamic acid itself does. The function of the dimethylamino group is to labilize the intermediate formed in the reaction, thus assisting deacylation intramolecularly. The dimethylamino group carries out this function by intramolecular general base catalysis. Nucleophilic catalysis is ruled out by the sizable deuterium oxide solvent isotope effect () found. General acid-hydroxide ion catalysis is ruled out by determination of the lack of reaction with azide ion, which does not possess a dissociable proton, with the intermediate in this reaction. The deuterium oxide solvent isotope effect on the azide ion reaction of the intermediate also rules out a general acid-hydroxide ion reaction. 相似文献
15.
On the basis of the calculated magnitude of the unidirectional flux through a gramicidin channel, it was predicted that a single conducting event should be sufficient to release trapped 22Na+ or 42K+ from phospholipid vesicles with a consequent apparent loss of ion selectivity. In support of this prediction, the introduction of gramicidin to the bathing solution of phospholipid vesicles containing trapped 22Na+ or 42K+ led to a release of vesicle contents which was consistent with the expectation that, for each gramicidin dimer present, the contents of approximately one vesicle are released. The predicted apparent loss of selectivity was also observed. Evidence was also presented suggesting some movement of gramicidin from vesicle to vesicle. The fluorescent intensity of gramicidin decreases with time when added to aqueous solutions at very low concentrations. It is proposed that this is a consequence of the extremely low solubility of gramicidin in water. On the basis of area per molecule calculations at the air-water interface, it was argued that the most likely conformation of gramicidin existing at the air-water interface, of those proposed in the literature, was that of ΠL,D6 helix. 相似文献
16.
Using inside-out thylakoid membranes, it has been shown that the oxidation of water and associated reduction of dichlorophenol indophenol is partially inhibited by low concentrations of cation chelators. This inhibition correlates with a removal of two manganese ions per Photosystem II reaction centre. The chelator-induced inhibition was completely reversed by the addition of low levels of Mn2+ () and higher levels of Mg2+ and Ca2+ (). Other cations were not effective, indicating that the ability to overcome the inhibition did not involve a general electrostatic screening process. The degree of inhibition by chelators was greater at lower light intensities and after treatment with glutaraldehyde. In the presence of glutaraldehyde the stimulatory effect of Mn2+ was lost, while pretreatment with Mn2+ prevented the glutaraldehyde effect. These results are discussed in terms of conformational changes of the electron donation chains involving cation- (preferentially Mn-) dependent coupling between the oxygen evolving and reaction-centre complexes of Photosystem II. 相似文献
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18.
The apparent Arrhenius energy of activation () of the water osmotic permeability () of the basolateral plasma cell membrane of isolated rabbit proximal straight tubules has been measured under control conditions and after addition of 2.5 mM of the sulfhydryl reagent, para-chloromercuribenzenesulfonic acid (pCMBS), of mersalyl and of dithiothreitol. (kcal/mol) was (controls) and (pCMBS), while decreased with pCMBS to of its control value. Mersalyl also decreased both in vitro and in vivo (using therapeutical doses). These actions of pCMBS and mersalyl were quickly reverted with 5 mM dithiothreitol and prevented by 0.1 M thiourea. for free viscous flow is 4.2 and greater than 10 for non-pore-containing lipid membranes. By analogy with these membranes and with red blood cells, where similar effects of pCMBS on are observed, it is concluded that cell membranes of the proximal tubule are pierced by aqueous pores which are reversibly shut by pCMBS. Part of the action of mercurial diuretics can be explained by their action on . 相似文献
19.
G.D. Holman 《生物化学与生物物理学报:生物膜》1980,599(1):202-213
Glucose transport in human erythrocytes is characterized by a marked asymmetry in the and values for entry and for exit. In addition, they show a high and a high for equilibrium exchange but low values for infinite cis and for infinite trans exit and entry. An allosteric pore model has been proposed to account for these characteristics. In this model, substrate-induced conformational changes destabilize the interfaces between protein subunits (the pore gates).Pores doubly occupied from inside destabilize the transport gates and result in high and high transport parameters. This effect is less marked when pores are doubly occupied from outside and therefore transport asymmetry results. 相似文献
20.
Using 1-6-12 empirical functions with a solvent-averaged electrostatic contribution and electrostatic potentials from CNDO-type wavefunctions, the development of specific interactions of ions visualized by the molecular electrostatic potential of PO4-group containing molecules was studied. Going from single molecules to monolayers made up of 37 head groups of phosphatidylcholine (PC) or phosphatidylethanolamine (PE) for quantum-chemical calculations, or of 23 head groups for empirical calculations we found decreasing potential minima. Only the inclusion of the screening effect of water, simulated by a distance dependent dielectric constant, ε(r), gives an explanation of stereospecific interactions of model membranes with ions. This finding can be compared with results of simulation calculations on water structure above a PE head group layer. 相似文献