共查询到20条相似文献,搜索用时 15 毫秒
1.
Nancy Sauberman Normand L. Fortier Grant Fairbanks Ronald J. OConnor L.Michael Snyder 《生物化学与生物物理学报:生物膜》1979,556(2):292-313
Significant alterations in the spectrin: band 3 and band 4.1a : band 4.1b ratios and an occasional decrease in the peak height of band 4.2 with respect to band 4.1 were found in electrophoretic patterns of red cell membranes from patients with hereditary xerocytosis. Electrophoretic comparison of whole cell, cytoplasm and membrane polypeptides implied that atypical partitioning at hemolysis could account for some, but not all, of the alterations seen in membrane patterns of xerocytes. A decrease in band 4.2 peak height as well as a variation in the profile of band 3 were produced in controls by specific manipulations of the electrophoresis protocol. Metabolic depletion of normal cells produced the type of alterations in bands 3 and 4.1 found in xerocyte membranes, whereas Heinz body production, addition of calcium to the hemolysis buffer and incubation of membranes in detergent under conditions designed to promote proteolysis did not. The presence of a higher peak height of band 4.1b with respect to that of band 4.1a in membranes of patients with various other red cell disorders correlated with an increase in the percentage of reticulocytes in peripheral circulation. The appearance of both band 3 and 4.1 abnormalities in the patterns of control cells which had been enriched in young cells by density gradient centrifugation suggested that these alterations in hemolytic disease are related to the predominance of young cells in the population. 相似文献
2.
Red cell membrane in hemolytic disease. Studies on variables affecting electrophoretic analysis 总被引:4,自引:0,他引:4
N Sauberman N L Fortier G Fairbanks R J O'Connor L M Snyder 《Biochimica et biophysica acta》1979,556(2):292-313
Significant alterations in the spectrin: band 3 and band 4.1a: band 4.1b ratios and an occasional decrease in the peak height of band 4.2 with respect to band 4.1 were found in electrophoretic patterns of red cell membranes from patients with hereditary xerocytosis. Electrophoretic comparison of whole cell, cytoplasm and membrane polypeptides implied that atypical partitioning at hemolysis could account for some, but not all, of the alterations seen in membrane patterns of xerocytes. A decrease in band 4.2 peak height as well as a variation in the profile of band 3 were produced in controls by specific manipulations of the electrophoresis protocol. Metabolic depletion of normal cells produced the type of alterations in bands 3 and 4.1 found in xerocyte membranes, whereas Heinz body production, addition of calcium to the hemolysis buffer and incubation of membranes in detergent under conditions designed to promote proteolysis did not. The presence of a higher peak height of band 4.1b with respect to that of band 4.1a in membranes of patients with various other red cell disorders correlated with an increase in the percentage of reticulocytes in peripheral circulation. The appearance of both band 3 and 4.1 abnormalities in the patterns of control cells which had been enriched in young cells by density gradient centrifugation suggested that these alterations in hemolytic disease are related to the predominance of young cells in the population. 相似文献
3.
The partitioning of fluorescence probes into intracellular organelles poses a major problem when fluorescence methods are applied to evaluate the fluidity properties of cell plasma membranes with intact cells. This work describes a method for resolution of fluidity parameters of the plasma membrane in intact cells labelled with the fluorescence polarization probe 1,6-diphenyl-1,3,5-hexatriene (DPH). The method is based on selective quenching, by nonradiative energy transfer, of the fluorescence emitted from the plasma membrane after tagging the cell with a suitable membrane impermeable electron acceptor. Such selective quenching is obtained by chemical binding of 2,4,6-trinitrobenzene sulfonate (TNBS), or by incorporation of glucosamine (BGA) to DPH-labelled cells. The procedures for determination of lipid fluidity in plasma membranes of intact cells by this method are simple and straightforward. 相似文献
4.
Takashi Watanabe Hajime Saito 《Biochimica et Biophysica Acta (BBA)/General Subjects》1980,633(1):77-86
Cytotoxicity and adsorption of pyocin S2 produced by Pseudomonas aeruginosa M47 (PAO 3047) to virally transformed mammalian cells, human malignant cells and normal cells in the same species were studied. Pyocin S2 inhibited the growth of not only tumor cells (XC, TSV-5, mKS-A TU-7, HeLa-S3 and AS-II cells) but also normal cells (BALB/3T3 and BHK 21 cells). The inhibitory effects on the cells increased with an increase of pyocin S2 activity. On the other hand, there were some tumor cells (155-4 T2 and HGC-27 cells) and normal cells (normal rat kidney and human embryo lung cells) which were resistant to pyocin S2. The pyosin S2 activity was neutralized by the cell membrane preparations from pyosin S2-sensitive cells, but not by those from pyocin-resistant cells. This neutralization ability was inhibited by high concentrations of D-galactose, neuraminic acid and completely destroyed by periodate and neuraminidase. The inhibition by the saccharides was concentration dependent. These results suggest that the toxicity of pyocin S2 to several mammalian cells is due to the presence of the binding site for pyocin S2 in the cell membrane and further, that the carbohydrate moiety, especially of D-galactose, and sialic acid, may play an important role as an initial binding site for pyocin S2. 相似文献
5.
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
6.
Haydn E. Edwards J.Kerry Thomas Gary R. Burleson Charles F. Kulpa 《生物化学与生物物理学报:生物膜》1976,448(3):451-459
The fluorescent probes pyrene, pyrene butyric acid and 1-naphthylamine have been used to investigate the changes that accompany in vitro transformation of a baby hamster kidney cell line using Rous sarcoma virus. The fluorescent probes which reside in the membrane were used to compare the changes in microviscosity and polarity of the membranes of normal cells with two transformed cell lines. The spectrofluorimetric data indicate that following transformation the probe 1-naphthylamine resides in a more polar environment. However, using the probe pyrene, the yield of excimer indicates decreased mobility of this probe in the membrane of transformed cells. The data also indicate differences between the two transformed cell lines. Laser photolysis was used to study the lifetime of the pyrne probes and the quenching of the pyrene fluorescence in the membrane by several different quenching molecules. The data indicate differences between the three cell lines and suggest that transformation decreases movement within the membrane. 相似文献
7.
When human red cells are treated with the mercurial sulfhydryl reagent, , osmotic water permeability is suppressed and only diffusional water permeability remains (Macey, R.I. and Farmer, R.E.L. (1970) Biochim. Biophys. Acta 211, 104–106). It has been suggested that the route for the remaining water permeation is by diffusion through the membrane lipids. However, after making allowance for the relative lipid area of the membrane, the water diffusion coefficient through lipid bilayers which contain cholesterol is too small by a factor of two or more. We have measured the permeability coefficient of normal human red cells by proton NMR and obtained a value of 4.0 · 10?3 cm · s?1, in good agreement with published values. In order to study permeation-through red cell lipids we have perturbed extracted red cell lipids with the lipophilic anesthetic, halothane, and found that halothane increases water permeability. The same concentration of halothane has no effect on the water permeability of human red cells, after maximal pCMBS inhibition. In order to compare halothane mobility in extracted red cell membrane lipids with that in red cell ghost membranes, we have studied halothane quenching of by equilibrium fluorescence and fluorescence lifetime methods. Since halothane mobility is similar in these two preparations, we have concluded that the primary route of water diffusion in pCMBS-treated red cells is not through membrane lipids, but rather through a membrane protein channel. 相似文献
8.
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
9.
Optimal conditions for activation of adenylate cyclase in membrane particles were studied. Enzyme activation with serotonin (5-hydroxytryptamine), NaF, and guanosine (GTPγS) was time- and temperature-dependent. Mg2+ was required for enzyme activation. Adenylate cyclase that was activated by NaF or GTPγS was gradually inhibited by while enzyme activated with serotonin and GTP responded faster to inhibition by the same sulfhydryl reagent. The enzyme responded in a similar fashion to a spin-labeled analog 3-(maleimidomethyl)-2,2,5,5-tetramethyl-1-pyrolidinyloxyl (i.e., nitroxide). Binding of the spin label was enhanced following enzyme activation by serotonin, NaF, or GTPγS in the presence of Mg2+. Activation of the enzyme was accompanied by an increase in the strong immobilization peaks in the EPR spectra. Both effects, the increase in binding and in the strong immobilization peaks, can be induced by Mg2+ alone. The results indicate that a general conformational change induced by Mg2+ may be essential for adenylate cyclase activation. 相似文献
10.
(1) (a penetrating SH- reagent) inactivated l-[14C]leucine entrance (binding and translocation) into Saccharomyces cerevisiae, the extent of inhibition depending on the time of preincubation with , concentration, the amino acid external and internal concentration, and the energization state of the yeast cells. With d-glucose-energized yeast, inhibited l-[14C]leucine entrance in all the assayed experimental conditions, but with starved yeast and low (0.1 mM) amino acid concentration, it did not inhibit l-[14C]leucine binding, except when the cells were preincubated with l-leucine. With the rho? respiratory-deficient mutant (energized cells), inhibited l[14C]leucine entrance as with the energized wild-type, though to a lesser extent. (2) Analysis of the effect as a function of l-[14C]leucine concentration showed a significant decrease of values of the high- (S1) and low- (S2) affinity amino acid transport systems, but values were not significantly modified. (3) When assayed in the presence of d-glucose, inhibition of d-glucose uptake and respiration contributed significantly to inactivation of l-[14C]leucine entrance. Pretreatment of yeast cells with 2,4-dinitrophenol enhanced the effect of l-[14C]leucine binding and translocation. (4) Bromoacetylsulfanilic acid and bromoacetylaminoisophthalic acid, two non-penetrating SH- reagents, did not inactivate l-[14C]leucine entrance, while , a slowly penetrating SH- reagent, inactivated it to a limited extent. When compared with the effect of , these negative results indicate that thiol groups of the l-[14C]leucine carrier were not exposed on the outer surface of the yeast cell permeability barrier. 相似文献
11.
Glycosylated compounds associated with the carbohydrate-rich tubular membrane system of the oxyntic cell were investigated. Two glycopeptide fractions, designated Peaks A and B, were isolated from pronase digests of bullfrog oxyntic cell microsomes. Molecular sieve chromatography and cellulose acetate electrophoresis revealed that, although somewhat heterogeneous, each peak was composed primarily of glycopeptides with similar molecular weights and net charge densities. Peak B glycopeptides had a mean molecular weight of about 6000 and contained 70% of the recovered carbohydrate in the following molar ratios: hexose, 1.00; , 0.71; fucose, 0.61; sialic acids, <0.03. Peak a glycopeptides were considerably larger (molecular weight approx. 100 000) and contained carbohydrates in molar ratios similar to those of Peak B. In both peaks galactose and , respectively, were the predominant hexose and amino sugar isomers.The glycolipid content of bullfrog oxyntic cell microsomes was assessed by qualitative and quantitative thin-layer chromatography. The most abundant glycolipids were monoglucosylceramides (0.098 mole/mole phospholipid) and monogalactosylceramides (0.046 mole/mole phospholipid). Small quantities of sulfatides and gangliosides were also present.A compilation of available data regarding the chemical composition of the microsomes revealed that these membranes resemble plasma membranes in having high molar ratios of cholesterol to phospholipid (approx. 1.0) and large quantities of carbohydrate (225 μg/mg protein). The possible significance of these compositional features in protecting the oxyntic cell is discussed. 相似文献
12.
Yukio Imanishi Hiroyuki Ohnishi Yutaka Hashimoto 《International journal of biological macromolecules》1981,3(2):97-104
The enantiomer selection in the nucleophilic addition reaction of optically active amines such as α-amino acid esters to phenylalanine and in as a solvent has been investigated. Stereoselectivity between the amines and the was found to change markedly according to the reaction conditions. This experimental finding is in contrast to the idea hitherto accepted that in the nucleophilic addition-type polymerization of α-amino acid the growing chain end reacts preferentially with one of the enantiomorphic having the same configuration, and indicates the importance of the investigation of stereoselectivity in the polymerization using suitable model reactions. Most acid esters reacted preferentially with , and this type of stereoselectivity increased with the of and with increasing bulkiness of the Cα substituent of α-amino acid esters (alanine < norleucine < leucine < valine). The relationship observed between the stereoselectivity and the structures of amines and was explained satisfactorily in terms of the transition state model in which the interaction of nitrogen and α-amino acid ester carbonyl as well as the interaction of carbonyl and α-amino acid ester nitrogen was taken into account. ethyl ester did not show enantiomer selectivity toward phenylalanine , but reacted preferentially with . for the reaction of proline ester with a transition-state model was proposed, which was different from the transition state model proposed for other α-amino acid esters. Some experiments were carried out to examine the transition-state models proposed. The implications of the present investigation in stereoselectivity in the nucleophilic addition-type polymerization of hitherto reported are discussed. 相似文献
13.
Oxidation of erythrocyte membrane SH-groups by diamide and tetrathionate induces cross-linking of spectrin (Haest, C.W.M., Kamp, D., Plasa, G. and Deuticke, B. (1977) Biochim. Biophys. Acta 469, 226–230). This cross-linking was now shown to go along with a concentration- and time-dependent enhancement of membrane permeability for hydrophilic nonelectrolytes and ions. The enhancement is specific for oxidative SH-group modifications, is reversible by reduction of the induced disulfides, can be suppressed by a very brief pre-treatment of the cells with low concentrations of and is strongly temperature-dependent. The pathway of the induced permeability discriminates nonelectrolytes on the basis of molecular size and exhibits a very low activation energy (). These findings are reconcilable with the formation of a somewhat inhomogeneous population of aqueous pores with radii probably . Estimated pore numbers vary with the size of the probe molecule. Assuming a diffusion coefficient as in bulk water within the pore, at least 20 pores per cell have to be postulated; more realistic lower diffusion coefficients increase that number. Alterations of the lipid domain by changes of cholesterol contents and insertion of hexanol or nonionic detergents alter the number or size of the pores. Since aggregation of skeletal and intrinsic membrane proteins also occurs after the SH-oxidation, in parallel to the formation of membrane leaks, one may consider (a) defects in the disturbed bilayer interface, (b) a mismatch between lipid and intrinsic proteins or (c) channels inbetween aggregated intrinsic proteins as structures forming the pores induced by diamide treatment. 相似文献
14.
Human red blood cells were treated with phospholipase C from Clostridium welchii. Lipase concentrations which produced <1% hemolysis and 10–15% hydrolysis of the membrane phospholipids reduced markedly () the accessibility of mambrane proteins to the external surface as measured by lactoperoxidase-catalyzed iodianation. 相似文献
15.
Yutaka Nishida Koichi Nabe Shigeki Yamada Ichiro Chibata 《Enzyme and microbial technology》1984,6(2):85-90
A procedure for production of () from its racemic mixture by enzymatic continuous hydrolysis of the l-enantiomer of () is described. For this hydrolysis, whole cells of Erwinia carotovora immobilized by κ-carrageenan gel were employed. The complete conversion of to was achieved by using a column packed with the immobilized cells. The half-life of the enzyme activity of the column was calculated to be ~100 days. Pure crystalline was readily isolated from the effluent with >87.5% yield. was easily racemized by incubating in ethanol containing 0.1 m NaOH for 30 min at 80°C, and used again for substrate. 相似文献
16.
Claudia Moore Merle L. Blank Ten-Ching Lee B. Benjamin Claude Piantadosi Fred Snyder 《Chemistry and physics of lipids》1978,21(3):175-178
In previous studies on the modification of polar head groups of membrane phospholipids with the unnatural base analog, , we reported an unidentified phospholipid in addition to in the various membrane fractions of rat liver. The structure of this phospholipid has now been identified as by nuclear magnetic resonance spectroscopy, and by chromatographic and enzymic analysis. In addition, we found that when rats were injected intraperitoneally with the , 19% of teh liver microsomal phospholipid was . 相似文献
17.
The DCCD-binding polypeptide is close to the F1 ATPase-binding site on the cytoplasmic surface of the cell membrane of Escherichia coli 总被引:2,自引:0,他引:2
Removal of the F1 ATPase from membrane vesicles of resulted in leakage of protons across the membrane through the FO portion of the ATPase complex. The leakage of protons was prevented by antiserum to the N,N′-dicyclohexylcarbodiimide (DCCD)-binding polypeptide in everted but not in “right-side out” membrane vesicles. The antiserum prevented the rebinding of F1 ATPase to F1-stripped everted membrane vesicles. It is concluded that in F1-depleted vesicles the DCCD-binding polypeptide is exposed on the cytoplasmic surface of the cell membrane at or close to the binding site of the F1 ATPase. 相似文献
18.
Graham K. Moore George A.F. Roberts 《International journal of biological macromolecules》1980,2(2):78-80
The gelation behaviour of solutions of chitosan on treatment with acyl anhydrides has been studied. The degree of required to initiate geletion has been found to depend on the molecular weight of the acyl anhydride and the concentration of chitosan, but to be independent of temperature. The energy of activation of has been determined for and . A mechanism has been proposed for the gelation process. 相似文献
19.
The saxitoxin-binding component of the excitable membrane sodium channel exhibits glycoprotein characteristics as evidenced by its specific interaction with various agarose-immobilized lectins. The detergent-solubilized saxitoxin-binding component interacts quantitatively with immobilized wheat germ agglutinin and concanavalin A and fractionally with immobilized Lens culinaris hemagglutinin and Ricinus communis agglutinin. These lectins preferentially bind and sialic acid (wheat germ agglutinin), mannose (concanavalin A and Lens cunilaris and galactose (Ricinus communis). Removal of terminal sialic acid residues by neuraminidase markedly decreases binding to immobilized wheat germ agglutinin but uncovers sites capable of interacting with lectins specific for galactose and . , an exoglycosidase has no effect on the binding of the channel protein to wheat germ agglutinin. Similarly, phospholipase C has no effect on binding of the solubilized toxin binding component to this lectin. Neither wheat germ agglutinin nor concanavalin A free in solution alters the number of toxin binding sites or their affinity for toxin. The sodium channel saxitoxin-binding component appears to be a glycoprotein containing terminal sialic acid residues and internal mannose, galactose, , and residues. The toxin binding site is spatially separated from the binding sites for the lectins studied. The effect of these sugar moieties must be considered when evaluating the biophysical parameters of the sodium channel. 相似文献
20.
A new polypeptide associated with RNA polymerase from Bacillus subtilis during late stages of vegetative growth 总被引:7,自引:0,他引:7
DNA-dependent RNA polymerase has been purified from at various stages of vegetative cell growth. Polymerase isolated from cultures approaching the end of the logarithmic growth phase was associated with a 60,000-dalton polypeptide and was only 10–20% as active as polymerase isolated from rapidly growing cells. Appearance of this new polypeptide and the change in template activity occur prior to stage 0 of sporulation. 相似文献