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1.
Conditions for the production of a complementary DNA sequence for use in studies of ribosomal RNA are described. . DNA polymerase I is used to transcribe highly purified 28S ribosomal RNA from rat liver. The reaction is sensitive to the tertiary structure of the rRNA template-primer. The complementary DNA hybridizes to its rRNA template with a of 0.02. The hybrid formed between 28S ribosomal RNA and complementary DNA has a Tm of 73°C. The probe reacts with total rat nuclear RNA with a of 1.0. 相似文献
2.
The presence of cytoplasmic poly(A) polymerase has been established in sea urchin eggs and four-cell embryos by subcellular fractionation and use of enucleate egg halves. ATP is the only ribonucleoside triphosphate incorporated. This incorporation is time dependent, contingent on input protein concentration, and immune to a variety of antimetabolites known to inhibit DNA-directed RNA synthesis. Both the unfertilized egg and the four-cell embryo cytoplasmic poly(A) polymerase activities display a preference for Mn2+. While oligo(A)4 is inactive as a primer, addition of and stimulates ATP incorporation. On a unit per milligram protein basis, the endogenous activity associated with cytoplasmic fractions obtained from nucleate and enucleate egg halves is 36 and 83% that obtained with the cytoplasmic fraction prepared from the unfertilized egg. In the presence of , both the nucleate and enucleate egg halves exhibit 81% of the activity associated with the unfertilized egg cytoplasmic fraction. The level of Mn2+ cytoplasmic poly(A) polymerase activity from the four-cell embryo is approximately 50% that of the unfertilized egg. This decrease does not appear to be due to either a postfertilization alteration in the subcellular localization of poly(A) polymerase or an increase in RNase activity. Supplementation with failed to restore the four-cell embryo cytoplasmic poly(A) polymerase potential to a level comparable to that of the unfertilized egg. Suppression of postfertilization protein synthesis by emetine, however, prevents this developmental decline in ATP incorporation thereby suggesting that postfertilization cytoplasmic poly(A) polymerase activity is subject to negative translational control. 相似文献
3.
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
4.
Differential polarized phase fluorometry was used to quantify the rotational rate () and limiting anisotropy () of the membrane probe diphenylhexatriene (DPH) in solvents and lipid vesicles exposed to hydrostatic pressures ranging from 1 bar to 2 kbar. These measurements reveal the effect of pressure on the phase-transition temperatures of the phosphatidylcholine vesicles, and the effects of pressure on order parameter of the acyl side-chain region of the membranes, the latter as indicated by . In addition to the well-known elevation of the transition temperature () with pressure, our results demonstrate that increased pressure restores the order of the bilayers to that representative of temperatures below the transition temperature. We also found that solvents which allowed free isotropic rotation of DPH at 1 bar no longer allowed free rotation when sufficiently compressed; moreover, the apparent DPH rotational rate increased with . Pressure studies using both DPH and the charged DPH analogue, trimethylammonium DPH (TMA-DPH) indicated that the of dipalmitoylphosphatidylcholine vesicles increased 23 K/kbar and an apparent volume change of 0.036 ml/mol lipid at the phase transition. Assuming, as has been proposed, that TMA-DPH is localized near the glycerol backbone region of the bilayers, these results indicate a similar temperature- and pressure-dependent phase transition in this region and the acyl side-chain region of the membrane. 相似文献
5.
Five species of cockroach were tested on a miniature treadmill at three velocities as O2 consumption () was measured: Gromphadorhina chopardi, Blaberus discoidalis, Eublaberus posticus, Byrsotria fumagata and Periplaneta americana. All cockroaches showed a classical aerobic response to running: increased rapidly from a resting rate to a steady-state on-response varied from under 30 s to 3 min. Recovery after exercise was rapid as well; off-response varied from under 30 s to 6 min. These times are faster or similar to mammalian values. varied directly with velocity as in running mammals, birds and reptiles. during steady-state running was only 4–12 times higher than at rest. Running is energetically much less costly per unit time than flying, but the cost of transport per unit distance is much more expensive for pedestrians. The minimal cost of transport (Mrun), the lowest necessary to transport a given mass a specific distance, is high in cockroaches due to their small size. The new data suggest that insects may be less economical than comparable sized vertebrates. 相似文献
6.
José Remacle 《生物化学与生物物理学报:生物膜》1980,597(3):564-576
The in vitro incorporation of cytochrome into purified plasma membranes was investigated by biochemical and immunological methods. Plasma membrane preparations incorporated three times less cytochrome than did microsomal preparations; 60% of this cytochrome could not be reduced by the NADH-cytochrome reductase and was considered as being bound to the plasma membrane. The morphological observations made after the immunochemical labeling of cytochrome clearly showed a good but asymmetrical distribution of the ferritin labeling: only the inner face of the plasma membrane incorporated cytochrome . These results are discussed with respect to theories which concern the subcellular membrane relationships in the cell. 相似文献
7.
G.Prem veer Reddy 《Biochemical and biophysical research communications》1982,109(3):908-915
Catalytic activity of thymidylate synthase, as measured , is tightly linked to S phase of the cell cycle in Chinese hamster embryo fibroblast cells. This activity, as measured , is found in all parts of the cell cycle. Thymidylate synthase activity in nuclear (karyoplast) extracts increased as the cells progressed from to S phase. This enzymatic activity in the nuclei of S phase cells is associated with the multienzyme complex (replitase) that also contained DNA polymerase and other enzymes of DNA replication and precursor synthesis. The degree of association of thymidylate synthase with replitase, which increased co-ordinately as the cells progressed from phase to S phase, coincided strongly with the level of activity of the enzyme. 相似文献
8.
J Remsen D Jerina H Yagi P Cerutti 《Biochemical and biophysical research communications》1977,74(3):934-940
The reaction of bacteriophage T7-DNA with the radioactive diastereomeric benzo(a)pyrene-diol-epoxides, (±) [, ]-7β,8α-dihydroxy-9α,10β-epoxy-7,8,9,10-tetrahydrobenzo(a)pyrene, and (±) [, ]-7β,8α-dihydroxy-9β,19β-epoxy-7,8,9,10-tetrahydrobenzo(1)pyrene, was investigated. Chromatographic analysis of digests of the DNA allowed the distinction of characteristic deoxynucleoside adduct peaks for the two benzo(a)pyrene-diol-epoxides. Our results, together with data from the literature, allow the identification of these adducts as mostly N2-(10-7β,8α,9α-trihydroxy-7,8,9,10-tetrahydrobenzo(a)pyreney1)deoxyguanosine and N2-(10-7β,8α,9β-trihydroxy-7,8,9,10-tetrahydrobenzo(a)pyreney1)deoxyguanosine, respectively. DNA-benzo(a)pyrene adducts with the same chromatographic properties were formed in mouse embryo fibroblasts upon treatment with benzo(a)pyrene. 相似文献
9.
Highly purified divalent and monovalent antibodies against cytochrome , anti- immunoglobulin G (IG) and anti- Fab', were used in elucidating the role of this cytochrome in the drug-oxidizing enzyme system of mouse liver microsomes. Anti- IG strongly inhibited not only NADH-supported but also NADPH-supported oxidation of 7-ethoxycoumarin and benzo(a)pyrene, but had no inhibitory action on the oxidation of aniline. Anti- Fab' also inhibited NADH-supported and NADPH-supported benzo(a)pyrene hydroxylation. These observations indicate an essential role of cytochrome in the transfer of electrons not only from NADH but also from NADPH to cytochrome P-450 in the microsomal oxidation of some drugs, but not of aniline. 相似文献
10.
A theoretical study of calcium entry in nerve terminals, with application to neurotransmitter release 总被引:4,自引:0,他引:4
In his study of a kin selection model for the evolution of workers' behavior in an incipiently social insect, Craig (1979) found that the haplodiploid mode of sex determination, combined with a female-biased sex ratio, cannot accelerate an evolutionary trend toward eusociality. This seems to contradict to Hamilton's (1964) theory. It is my intention to prove that Craig's result is due to the dependence of relative reproductive success in each sex on the sex ratio of a population. The oviposition of unfertilized eggs by workers is indispensable, in primitive stages in the evolution of eusociality, for maintaining the relative reproductive success of a female. Assuming that workers control the sex ratios, we can distinguish the following three evolutionary phases in accordance with the ratio (the ratio of the number of the queen's second brood to that of the progeny laid by workers derived from the same nest). During Phase 1 in which < , the reproductive successes of a male and a female are equal, and Hamilton's rule holds. In Phase 2 in which , the queen produces females, and workers oviposit males. As increases, the sex ratio in the whole population becomes female-biased and relative reproductive success of a male increases. In Phase 3 in which N2/N10 >, the queen lays some male eggs in addition to female eggs. The sociality threshold ()crit the minimum value of the benefit/cost ratio leading to the evolution of altruism, is on Phase 1. It rises threefold as N2/N10 increases in Phase 2, and, in Phase 3, it is twice as high as that in a diploid species. The anatomical and physiological characteristics of workers must have been so developed that they are efficient in helping activities after the beginning of eusociality. The evolutionary process before the beginning of helping behavior is also discussed. The egg laying by unmated females, which stay their mother's nest, seems to have been an important preadaptation for the evolution of eusociality in Hymenoptera. 相似文献
11.
The effect of prostaglandins E1 and E2 on the human erythrocyte as monitored by spin labels 总被引:8,自引:0,他引:8
P G Kury P W Ramwell H M McConnell 《Biochemical and biophysical research communications》1974,56(2):478-483
The effects of the prostaglandins PGE1 and PGE2 on the deformability of the human erythrocyte were studied using spin-labeled erythrocytes. Two magnetic resonance parameters were measured: (1) The orientation relaxation time, , for the erythrocyte, and (2) the order parameter, S, for a fatty acid spin label bound to the membrane. Prostaglandins PGE1 and PGE2 exhibited opposite effects on both and S. PGE2 made the cell less deformable (increases of and S) and PGE1 made the erythrocyte more deformable (decrease of and S). 相似文献
12.
J P Henry F Hirata O Hayaishi 《Biochemical and biophysical research communications》1978,81(4):1091-1099
Superoxide anion can serve a reducing agent for tyramine hydroxylation by dopamine-β-hydroxylase. Stable solutions were obtained by dissolving KO2 in dry dimethylsulfoxide and infused into buffered solutions of tyramine and dopamine-β-hydroxylase at constant rate. The reaction requires molecular oxygen, but differs from the ascorbate dependent hydroxylation in its alkaline pH optimum value (pH 7.5) and its low rate (9 nmol octopamine formed/min/mg of protein). In absence of tyramine does not produce a stable reduced form of the enzyme. 相似文献
13.
Zoltán Spolarics Jozsef Mandl Raymund Machovich Patrick Lambin Tamás Garzó Ferenc Antoni István Horváth 《Biochimica et Biophysica Acta (BBA)/Molecular Cell Research》1985,845(3):389-395
125I-labelled complexed with thrombin or plasmin bound to hepatocytes in a concentration-and time-dependent manner. The apparent values calculated from displacement experiments were 7.9 · 10?8 M for and 8.5 · 10?8 M for . Association of these complexes was only partially reversible; after a 180 min incubation period, 50–60% of the bound radioactivity was internalized by the cells. itself bound also to hepatocytes, but the affinity of the complexes was higher than that of the inhibitor alone, and was not internalized, either. 125I-labelled thrombin or plasmin bound to hepatocytes as well. These bindings were also concentration-dependent and could be decreased with an excess of unlabelled ligands. Binding rates and amounts of the bound proteinases were higher than those of their complexes. The complex competed with the complex in binding to hepatocytes, whereas there was no competition between these complexes and the antithrombin III-thrombin complex. These results suggest that the binding sites of hepatocytes for and antithrombin III-proteinase complexes are different. 相似文献
14.
The distribution of activity was studied in cell fractions prepared from homogenates of rat liver. The level of mitochondrial contamination in the microsomal fraction depended on the fractionation procedure and on the method of homogenization. With proper care, microsomes with undetectable mitochondrial contamination could be prepared. These microsomes had no detectable activity. Approximately 85 % of the total activity of the post 6000 x g · min supernatant was recovered in the mitochondrialfraction. The properties of this mitochondrial were not distinguishable from those of the various microsomal previously described by other investigators. 相似文献
15.
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
16.
D F Kimball L Peterson D J McLoughlin R G Wolfe 《Archives of biochemistry and biophysics》1979,195(1):66-73
Initial rate, product inhibition, and isotope rate kinetic studies of pig heart mitochondrial and supernatant malate dehydrogenases, acting upon the nonphysiological substrates, meso-tartrate and 2-keto-3-hydroxysuccinate, are reported. The measured spontaneous keto-enol equilibrium for 2-keto-3-hydroxysuccinate in 0.05 m Tris-acetate (pH 8.0) at 25 °C favors the enol form, dihydroxyfumarate, with an apparent equilibrium constant of 0.036. The enzyme-catalyzed reaction favors meso-tartrate with an apparent equilibrium constant of 1.25 × 10?6, M?1 at pH 8.0. The mechanism apparently remains ordered bi bi for both enzymes when these nonphysiological substrates are used, and the chemical-converting hydride transfer step becomes more rate limiting for both enzymes. This conclusion is supported by and values of 2.6 and 3.1, respectively, for the mitochondrial enzyme and 1.9 and 2.9, respectively, for the supernatant enzyme. 相似文献
17.
M.Alfonsina Desiderio Angela Sessa Antonio Perin 《Biochimica et Biophysica Acta (BBA)/Molecular Cell Research》1985,845(3):463-468
The administration of preferential adrenergic receptor antagonists to uninephrectomized rats revealed the mediation in diamine oxidase activity increase that occurs in the remaining kidney undergoing compensatory hypertrophy. In fact, or , but not , , or this enzyme enhancement. Further studies with adrenoceptor agonists, such as epinephrine (, , , ), isoproterenol (, ) or terbutaline () showed that also in normal rat kidney diamine oxidase activity is under the control of through a mechanism that involves new synthesis of mRNA and protein. Theophylline, an inhibitor of phosphodiesterase, or forskolin, an activator of adenyl cyclase, increased diamine oxidase activity as does epinephrine or nephrectomy. Thus, catecholamine-triggered coupled to adenyl cyclase are involved in the regulation of diamine oxidase activity in normal and hypertrophic rat kidney. 相似文献
18.
Interaction of lymphocytes and macrophage cell line cells (M1 cells). I. Functional maturation and appearance of Fc receptors im M1 cells 总被引:2,自引:0,他引:2
M1 cells, which are cell line cells established from myeloid leukemia cells of the SL strain mouse, can differentiate from blast cells () to mature macrophages () within 48 hr, when they are cultured with conditioned medium (CM) obtained from murine embryonic fibroblasts. While cells have no phagocytic activity nor Fc receptor (FcR), cells possess both characteristics. The appearance of FcR is temperature-dependent and inhibited by a metabolic inhibitor, cycloheximide. FcR on cells is resistant to trypsin and pronase. cells improve the viability of macrophage-depleted SL splenic lymphocytes and restore the in vitro secondary plaque forming cell response of macrophage-depleted spleen cells to particulate and soluble antigens. cells lack this macrophage-substituting capacity. Mm1 cells, mutant cells from M1 cells, having FcR and higher phagocytic activity than cells, are also devoid of this capacity. 相似文献
19.
Nancy B. Schwartz Victoria Ostrowski Kenneth S. Brown Robert M. Pratt 《Biochemical and biophysical research communications》1978,82(1):173-178
Activity levels of sulfotransferases, requisite for the sulfation of chondroitin sulfate proteoglycan, were measured in cell-free homogenates prepared from neonatal epiphyseal cartilage of normal C57B1/6J or homozygous brachymorphic mice. In the presence of [35S]-PAPS only or [35S]-PAPS plus an exogenous sulfate acceptor, comparable amounts of were incorporated into chondroitin sulfate by the normal and mutant types of cartilage. In contrast, the mutant cartilage catalyzed the conversion of only 30% of the into chondroitin sulfate as compared to normal mouse cartilage when synthesis was initiated from ATP and H235SO4. These results suggest that the production of an undersulfated proteoglycan which has previously been reported in brachymorphic mice (Orkin, R.W. . (1976) Devel. Biol. , 82–94) may result from a defect in the synthesis of the sulfate donor PAPS. 相似文献
20.
Purified cytochrome from bovine adrenocortical mitochondria was incorporated into liposomes by the cholate-dilution method utilizing either dialysis or Sephadex gel filtration. Among synthetic phospholipids tested, dioleoylglycerophosphocholine showed the best stability during the incorporation of into liposomes. A maximum amount of heme was incorporated into liposomes at a molar ratio of phospholipid to the cytochrome of approx. 200. When was incorporated into the dioleoylglycerophosphocholine liposomes by the cholate-filtration method, the showed two major populations on the elution pattern of the Sepharose 4B gel filtration, and were seen at a diameter of 200–600 Å and its aggregated forms. When the cytochrome was incorporated into dioleoylglycerophosphocholine liposomes or cholesterol-free adrenocortical mitochondrial liposomes, was less stable than in aqueous solution. Cholesterol or adrenodoxin markedly stabilized the liposomal . Liposomal required cholesterol for its optimum reduction with adrenodoxin, adrenodoxin reductase, and NADPH in the presence of CO. About 70% of the total heme in the dioleoylglycerophosphocholine liposomes was reduced by the enzymatic reduction in the presence of cholesterol, indicating that 70% of the total molecules are exposed to the surface of the outer monolayer. In order to see the location of the heme in membrane, the dioleoylglycerophosphocholine-liposomal was subjected to treatment. This reagent destroyed the liposomal . These results suggest that the heme is located in the proximity of the reacting sites which are exposed to the surface, or located on the vincinity of polar heads of the membrane. 相似文献