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71.
PPi driven ATP synthesis has been reconstituted in a liposomal system containing the membrane-bound energy-linked PPiase and coupling factor complex, both highly purified from Rhodospirillum rubrum. This energy converting model system was made by mixing both enzyme preparations with an aqueous suspension of sonicated soybean phospholipids and subjecting to a freeze-thaw procedure. In the presence of ADP, Mg2+, Pi and PPi the system catalyzed phosphorylation by up to 25 nmol ATP formed X mg protein-1 X min-1, at 20 degrees C, which was sensitive to uncouplers and inhibitors of phosphorylation such as oligomycin, efrapeptin and N,N'-dicyclohexylcarbodiimide. 相似文献
72.
The temperature dependence of the kinetics of the binding of ATP to myosin subfragment-1 was studied by an ATP chase technique in a rapid-flow-quench apparatus: (formula; see text) A temperature range of 30 degrees C to -15 degrees C was obtained with ethylene glycol as antifreeze. The Arrhenius plot of k2 is discontinuous with a jump at 12 degrees C. Above the jump delta H+ = 9.5 kcal/mol, below delta H+ = 28.5 kcal/mol. Few such Arrhenius plots are recorded in the literature but they are predicted from theory. Thus, we explain our results as a phase change of the subfragment 1-ATP system at 12 degrees C. This is in agreement with certain structural studies. 相似文献
73.
Calcium-dependent activation and deactivation of rod outer segment phosphodiesterase is calmodulin-independent 总被引:2,自引:0,他引:2
ATP-dependent activation and deactivation of retinal rod outer segment phosphodiesterase is affected by calcium [Kawamura, S. and Bownds, M. D., J. Gen. Physiol. 77:571-591(1981)]. Our data demonstrate that although calmodulin has been found in rod outer segments [Liu, Y. P. and Schwartz, H., Biochim. Biophys. Acta 526:186-193(1978); Kohnken, R. E. et al, J. Biol. Chem. 256:12517-12522(1981)], this protein is not involved in calcium-dependent phosphodiesterase activation at light levels at which calcium clearly affects this enzyme's activity. Furthermore, calmodulin does not mediate the calcium-dependent deactivation of phosphodiesterase. 相似文献
74.
John A. Jacquez 《生物化学与生物物理学报:生物膜》1983,727(2):367-378
Depletion of energy stores of human red cells decreases the maximum transport capacity, , for glucose transport to a value one-third or less of that found in red cells from freshly drawn blood. There is no change in . Hemolysis and resealing of red cells with ATP or ADP reverses the decrease in . The maximum effect occurs at concentrations of ATP in the normal range for red cells, however, there is little effect from ADP concentrations in its normal range in freshly drawn red cells. Hemolysis and resealing with ATP gives an increase in and an increase in differential labeling by photolytic labeling with tritiated cytochalasin B. Most of the activation is lost after a second hemolysis-reseal without ATP but about 25% of the activation remains. 相似文献
75.
Delocalized chemiosmotic coupling of oxidative phosphorylation requires that a single-value correlation exists between the extent of and the kinetic parameters of respiration and ATP synthesis. This expectation was tested experimentally in nigericin-treated plant mitochondria in single combined experiments, in which simultaneously respiration (in State 3 and in State 4) was measured polarographically, FΔψ (which under these conditions was equivalent to ) was evaluated potentiometrically from the uptake of tetraphenylphosphonium+ and the rate of phosphorylation was estimated from the transient depolarization of mitochondria during State 4-State 3-State 4 transitions. The steady-state rates of the different biochemical reactions were progressively inhibited by specific inhibitors active with different modalities on various steps of the energy-transducing process: succinate respiration was inhibited competitively with malonate or noncompetitively with antimycin A, or by limiting the rate of transport into the mitochondria of the respiratory substrate with phenylsuccinate; was dissipated by uncoupling with increasing concentrations of valinomycin; ADP phosphorylation was limited with oligomycin. The results indicate generally that when the rate of respiratory electron flow is decreased, a parallel inhibition of the rate of phosphorylation is also observed, while very limited effects can be detected on the extent of . This behavior is in marked contrast to the effect of uncoupling where the decreased rate of ATP synthesis is clearly due to energy limitation. Extending previous observations in bacterial photosynthesis and in respiration by animal mitochondria and submitochondrial particles the results indicate, therefore, that respiration tightly controls the rate of ATP synthesis, with a mechanism largely independent of . These data cannot be reconciled with a delocalized chemiosmotic coupling model. 相似文献
76.
The secretory system of intact chromaffin cells is not accessible to direct chemical manipulation because of the selective permeability of the plasmalemma. We have devised a simple procedure for chemically "skinning" (permeabilizing) cultured adrenal medullary chromaffin cells by brief exposure to the detergent saponin. This procedure disrupts the continuity of the plasmalemma, thus allowing us to bypass those aspects of the secretory process controlled by the cell membrane and giving direct access to exogenous substances to the cellular secretory machinery. We report here that the skinned cells retain a fully competent secretory mechanism dependent only on exogenous calcium and MgATP. Saponin treatment had no significant effect on the total catecholamine content of the cells. Secretion could be initiated by either MgATP or calcium as long as the other was present in the medium. Catecholamine and dopamine-beta-hydroxylase release by the skinned cells was dependent on the calcium concentration of the medium. The ratio of secreted catecholamine and enzyme was similar to that of the cells, indicating that secretion occurred by an exocytotic mechanism. About half the total cellular content of the cytoplasmic enzyme lactic dehydrogenase was released during the permeabilization process and subsequent incubations, indicating plasmalemma permeability to molecules as large as protein. Calcium-induced secretion was unaffected by several drugs known to affect catecholamines and granule function. Saponin treatment of chromaffin cells in culture appears to be a simple means for allowing access to exogenous substances to the cells' secretory machinery. Therefore, it offers the opportunity to use chemical treatments, and perhaps specific antibodies to cellular components, to determine the role of these elements in the secretory process. These techniques should also be applicable to other cells known to secrete by an exocytotic mechanism. 相似文献
77.
Acetylcholine Releases ATP from Varicosities Isolated from Guinea Pig Myenteric Plexus 总被引:2,自引:1,他引:1
The effects of cholinergic agonists and antagonists on the release of ATP from isolated myenteric varicosities were studied using a firefly luciferin-luciferase technique. In a previous study, acetylcholine and nicotine released ATP from isolated myenteric varicosities, whereas the muscarinic agonist bethanechol did not. In the present study, release of ATP by acetylcholine was shown to be Ca2+ dependent. d-Tubocurarine competitively antagonized the release of ATP by either acetylcholine or nicotine. Maximal release of ATP by acetylcholine (10(-3) M) was approximately 24% that observed with the depolarizing drug veratridine (5 X 10(-5) M), suggesting either that not all of the varicosities capable of releasing ATP possess nicotinic receptors or that acetylcholine does not depolarize the varicosities to the degree that veratridine does. Tetrodotoxin slightly but significantly reduced ATP release induced by acetylcholine or nicotine, indicating some involvement of Na+ channels in the release process. Finally, 6-hydroxydopamine pretreatment produced a 48% reduction in the acetylcholine-evoked release of ATP, suggesting that much, but possibly not all, of the ATP release occurs from noradrenergic varicosities present in the preparation. 相似文献
78.
A chemiosmotic molecular mechanism for proton-translocating adenosine triphosphatases 总被引:16,自引:0,他引:16
P Mitchell 《FEBS letters》1974,43(2):189-194
79.
The proton-translocating ATPase of Escherichia coli 总被引:17,自引:0,他引:17
80.
Rat liver cytosol binds 3H-cAMP and 3H-DBcAMP . Fractionation of bound radioactivity by DEAE-Sephadex chromatography shows that 3H-cAMP is associated with a different cytosolic protein than is 3H-DBcAMP. The pI's of the cAMP-protein and the 3H-DBcAMP-protein complexes are 6.7 and 3.9, respectively. Competition studies between 3H-cAMP and its structural analogues have shown the following order of effectiveness in competing for binding sites in rat liver cytosol: cAMP > N6-MBcAMP > O2′-MBcAMP. No inhibition of 3H-cAMP binding was observed with 5′-AMP, adenosine, cGMP or DBcAMP. binding experiments with rat serum has shown that only 3H-DBcAMP binds to any significant extent. 相似文献