Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.6.1.3 (ATPase)
 65,361 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We report a procedure for the large-scale purification of the Escherichia coli Rep protein, a helicase that is involved in the replication of the E. coli chromosome as well as a number of single-stranded bacteriophages. The procedure starts with E. coli cells harboring an overproducing plasmid, pRepO, in which the E. coli rep gene is under transcriptional control of the inducible lambda PL promoter (Colasanti, J., and Denhardt, D. T. (1987) Mol. Gen. Genet. 209, 382-390). The purification procedure results in greater than 98% pure Rep protein, which is free of contaminating nuclease activity, with yields of 40-50 mg of Rep protein/50 g of induced MZ-1/pRepO cells. We also show that cell death occurs upon inducing such a large overproduction of the E. coli Rep protein in MZ-1/pRepO. The Rep protein purified by this procedure has high specific single-stranded DNA-dependent ATPase activity, as well as helicase activity, with an apparent 3' to 5' directionality. The extinction coefficient of purified E. coli Rep protein is epsilon 280 = 1.16 +/- 0.04 ml mg-1 cm-1 (8.47 +/- 0.28 X 10(4) M-1 cm-1) in 10 mM Tris (pH 7.5), 20% (v/v) glycerol, 0.10 M NaCl at 25 degrees C. The solubility properties of the purified Rep protein have been examined as a function of glycerol, NaCl, MgCl2, ATP, and ADP concentrations at 25 and 37 degrees C (pH 7.5). Rep protein solubility decreases significantly with decreasing concentrations of glycerol and monovalent salt and increasing temperature; however, the presence of 1.5 mM ATP or ADP or MgCl2 at low NaCl concentrations increases the solubility. At 4 degrees C, in the presence of 20% glycerol and greater than or equal to 50 mM NaCl, the free Rep protein exists as a stable monomer under all conditions examined (+/- ATP and +/- MgCl2). The single-stranded DNA-dependent ATPase activity decreases with increasing glycerol concentration, such that in 25% (v/v) glycerol it has approximately 40% of its activity as compared to solutions that contain no glycerol. The dependence of the single-stranded DNA-dependent ATPase activity on salt concentration for a series of monovalent salts indicates the presence of both cation and anion effects, with decreasing activity in the order glutamate greater than acetate greater than chloride. The ability to obtain highly purified E. coli Rep protein in large quantities with relative ease will greatly facilitate physical characterizations of the protein and its interactions with DNA.
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PMID:Large-scale purification and characterization of the Escherichia coli rep gene product. 252 89

1. The toxic plant sesquiterpene lactones, helenalin, hymenoxon, mexicanin-E, tenulin, dihydrogriesenin, and psilotropin which were isolated from Helenium, Hymenoxys, and Geigeria spp markedly inhibited "state 3" respiration in mouse hepatic mitochondria. With the exception of dihydrogriesenin and psilotropin, all the other sesquiterpene lactones stimulated "state 4" respiration. 2. The sesquiterpene lactones also stimulated ATPase activity in the presence of Mg2+ ions and caused mitochondrial swelling in buffer solutions containing magnesium, sodium, ammonium and potassium chloride salts. 3. The number of alkylating sites present in a sesquiterpene lactone appears to be related to the inhibitory activity on mitochondrial oxidative phosphorylation, however, the structure-activity relationship of these lactones is not clear at present. 4. Mitochondria prepared from the liver of ethoxyquin hydrochloride-fed mice were less susceptible to sesquiterpene lactone-mediated inhibition of mitochondrial oxidative phosphorylation.
Gen Pharmacol 1989
PMID:Effects of sesquiterpene lactones on mitochondrial oxidative phosphorylation. 253 10

1. Direct stimulation evoked twitches in mouse diaphragm muscles in presence of 10 microM D-tubocurarine in vitro. Effects of ouabain and their dependence on K+ were examined on the twitch responses and action potentials in the presence and absence of twitch potentiators. 2. Ouabain inhibited twitch contractions only in the presence of veratridine, aconitine and monensin while it had no inhibitory effect on control twitches. The interactions between ouabain and these twitch potentiators depended on the presence of external K+, except in the case of monensin. 3. Removal of Ca2+ from a bathing solution accelerated the potentiating effect of veratridine and the antagonizing effect of ouabain. 4. Caffeine further potentiated the twitches which had been attenuated by ouabain combined with veratridine. 5. Ouabain combined with veratridine consistently decreased resting membrane potentials, action potentials and overshoot potentials and prolonged time to peak of and duration of the muscle action potentials. 6. Tetraethylammonium, 4-aminopyridine, and caffeine produced twitch potentiation which was insensitive to ouabain or the removal of K+. 7. These results suggest that twitch contractions in the presence of activators of sodium channels link with activation of Na+-K+-ATPase. Accumulation of Na+ inside the muscle fibres may uncouple the excitation-contraction system. 8. This uncoupling may not include the caffeine-sensitive process that controls the release of Ca2+ from the sarcoplasmic reticulum. Na+ accumulation may decrease transmembraneous gradient of this ions, thereby causing a reduction in excitation coupled with twitch contraction.
Gen Pharmacol 1989
PMID:The influence of ouabain on twitch contractions in the presence of veratridine. 254 58

Myosin was isolated from pig atrial and ventricular myocardium during postnatal development and Ca2+-ATPase was determined and myosin light chains were analysed by electrophoresis in sodium dodecylsulfate polyacrylamide gel. During ontogenesis ATPase activity of ventricular myosin remains virtually unchanged, whereas that of atrial myosin increases. The patterns of myosin light chains of atrial and ventricular myosin differ from each other, but the individual pattern remains unchanged during the development.
Gen Physiol Biophys 1989 Feb
PMID:Ontogenic differentiation of pig atrial and ventricular myosin. 254 80

In order to examine the dynamics of ion regulation, osmoregulation, and plasma calcitonin during the parr-smolt transformation (smoltification), blood and gill tissue were collected from yearling coho salmon, Oncorhynchus kisutch, from February to October. Fish were kept in fresh water (FW) throughout this period. In addition, fish were exposed to seawater (SW) at the peak of smoltification in mid-April, and samples from these fish were collected until July. Plasma osmolality, gill Na+,K+-ATPase activity, plasma levels of calcitonin, and free and total calcium and magnesium were measured. SW adaptability of FW fish was assessed throughout the study by measurements of plasma osmolality following a 24-hr exposure to seawater. The greatest hypoosmoregulatory ability occurred in April-May, although SW-adapted fish had higher plasma osmolality than FW-adapted fish at all times. Gill Na+,K+-ATPase activity in FW-adapted fish increased from April to June and increased rapidly following exposure of fish to SW, and remained elevated in SW-adapted fish. Free plasma calcium and magnesium levels increased following SW exposure, but returned to prior levels within 1 week. Netting and confinement stress during sampling caused an increase in plasma osmolality and free calcium and magnesium levels in both FW- and SW-adapted fish. Changes in hypoosmoregulatory ability during smoltification and SW adaptation were correlated with changes in gill Na+,K+-ATPase activity. A sharp transitory peak in plasma calcitonin levels occurred early in smoltification (March) and in SW-adapted fish in June. Plasma calcitonin levels gradually increased in FW-adapted fish during the period of desmoltification. However, no change in plasma calcitonin levels occurred during SW-induced hypercalcemia, suggesting that the hormone does not play a major role in short-term plasma calcium regulation in coho salmon.
Gen Comp Endocrinol 1989 Jun
PMID:Smoltification and seawater adaptation in coho salmon (Oncorhynchus kisutch): plasma calcium regulation, osmoregulation, and calcitonin. 254 13

To obtain more information on the role of prolactin and growth hormone during the parr-smolt transformation of Atlantic salmon, a population of fish in fresh water was sampled from January to June during two consecutive years. Gill Na+,K+-ATPase activity increased steadily during smoltification and a plasma thyroxine peak was observed 2-3 weeks before the gill Na+,K+-ATPase peak. On the basis of these two parameters, smoltification was considered complete in our populations in April 1985 and May 1986. Two peaks in plasma growth hormone levels occurred in 1986, one in mid-April and the second in mid-May. In both cases, these peaks coincided with a peak in plasma triiodothyronine and preceded the thyroxine peak by 1-2 weeks. Moreover, the second peak which lasted for 1 month coincided with maximal gill Na+,K+-ATPase activity. A decrease in plasma prolactin levels was observed during smoltification of Atlantic salmon in 2 consecutive years. During this period of decreasing and low plasma prolactin levels, gill Na+,K+-ATPase activity increased to its highest values. Atlantic salmon smolts were also directly transferred into seawater. After 2 days or more in seawater, plasma prolactin levels were not significantly different from those on Day 0, whereas in fresh water they showed large fluctuations. All these data indicate that growth hormone may play an important role in the development of hypoosmoregulatory activity. Increased hypoosmoregulatory ability also appears to be associated with low prolactin levels.
Gen Comp Endocrinol 1989 Jun
PMID:Smoltification and seawater adaptation in Atlantic salmon (Salmo salar): plasma prolactin, growth hormone, and thyroid hormones. 254 14

Stimulation by serum of cell proliferation in G1-arrested culture of Chinese hamster ovary cells CHO-K1 was accompanied by an early (during the first minutes) and delayed (2-10 h) activation of Na+,K+-ATPase and an increase in cell K+ content from 0.5-0.6 to 0.7-0.8 mmol per gram protein. Isoproterenol acted synergistically with serum in eliciting both early and delayed changes in K+ transport and in stimulating G1----S transition. Isoproterenol alone (without serum) induced a transient increase in K+ influx via Na+,K+-ATPase without changing the cell K+ content or having any mitogenic effect. Theophylline enhanced the serum-induced early activation of Na+,K+-ATPase but inhibited both the delayed increase in cell K+ and the G1----S transition. Early serum-induced increase in K+ transport was not affected by cycloheximide, whereas net accumulation of cell K+ was abolished by the drug. It is concluded that the early and the delayed activation of Na+,K+-ATPase induced by mitogens can be dissociated; the early ionic response is related to the primary transduction of membrane signal, whereas the delayed modulation of ion transport via Na+,K+-ATPase has another function and is associated with cell growth.
Gen Physiol Biophys 1989 Jun
PMID:Early and delayed changes in potassium transport during the initiation of cell proliferation in CHO culture. 254 16

The plasma-membrane ATPase of Saccharomyces cerevisiae is a proton pump whose activity, essential fro proliferation, is subject to regulation by nutritional signals. The previous finding that the CDC25 gene product is required for the glucose-induced H+-ATPase activation suggested that H+-ATPase activity is regulated by cAMP. Analysis of starvation-induced inactivation and glucose-induced activation of the H+-ATPase in mutants affected in activity of the RAS proteins, adenylyl cyclase or cAMP-dependent protein kinase showed that nutritional regulation of H+-ATPase activity does not depend directly on any of these factors. We conclude that adenlyl cyclase does not mediate all nutritional responses. This also indicates that the specific CDC25 requirement for the glucose-induced activation of the H+-ATPase identifies a new function for the CDC25 gene product, a function that appears to be independent of CDC25-mediated modulation of the RAS/adenylyl cyclase/cAMP pathway.
J Gen Microbiol 1989 Jun
PMID:cAMP- and RAS-independent nutritional regulation of plasma-membrane H+-ATPase activity in Saccharomyces cerevisiae. 255 50

Phosphorylation of red cell membranes at ambient temperatures with micromolar [32P]ATP in the presence of Na ions produced phosphoenzyme that was dephosphorylated rapidly upon the addition of ADP or K ions. However, as first observed by Blostein (1968, J. Biol. Chem., 243:1957), the phosphoenzyme formed at 0 degrees C under otherwise identical conditions was insensitive to the addition of K ions but was dephosphorylated rapidly by ADP. This suggested that the conformational transition from ADP-sensitive, K-insensitive Na pump phosphoenzyme (E1 approximately P) to K-sensitive, ADP-insensitive phosphoenzyme (E2P) is blocked at 0 degrees C. Since the ATP:ADP exchange reaction is a partial reaction of the overall enzyme cycle dependent upon the steady state level of E1 approximately P that is regulated by [Na], we examined the effects of temperature on the curve relating [Na] to ouabain-sensitive ATP:ADP exchange. The characteristic triphasic curve seen at higher temperatures when [Na] was between 0.5 and 100 mM was not obtained at 0 degrees C. Simple saturation was observed instead with a K0.5 for Na of approximately 1 mM. The effect of increasing temperature on the ATP:ADP exchange at fixed (150 mM) Na was compared with the effect of increasing temperature on (Na + K)-ATPase activity of the same membrane preparation. It was observed that (a) at 0 degrees C, there was significant ouabain-sensitive ATP:ADP exchange activity, (b) at 0 degrees C, ouabain-sensitive (Na + K)-ATPase activity was virtually absent, and (c) in the temperature range 5-37 degrees C, there was an approximately 300-fold increase in (Na + K)-ATPase activity with only a 9-fold increase in the ATP:ADP exchange. These observations are in keeping with the suggestion that the E1 approximately P----E2P transition of the Na pump in human red cell membranes is blocked at 0 degrees C. Previous work has shown that the inhibitory effect of Na ions and the low-affinity stimulation by Na of the rate of ATP:ADP exchange occur at the extracellular surface of the Na pump. The absence of both of these effects at 0 degrees C, where E1 approximately P is maximal, supports the idea that external Na acts through sites on the E2P form of the phosphoenzyme.
J Gen Physiol 1985 Jan
PMID:Temperature effects on sodium pump phosphoenzyme distribution in human red blood cells. 257 48

The in vitro action of SP and theophylline on the activities of (Na K) and (Ca Mg) dependent adenosine-5'-triphosphates (ATPase) of crude synaptosomal membranes (SM) of four different areas of rat brain was investigated. The highest activity of (Ca Mg)ATPase was observed in SM from hippocampus. In thalamus with hypothalamus region the highest activity of (Na K)ATPase and the lowest activity of (Ca Mg)ATPase occurred. It was found, that 10 mumol of SP stimulating "alternatively" both (Na K) and (Ca Mg)ATPase activities from different regions may constitute a regulating agent for ionic transport in CNS of rat. Theophylline on concentration of 5 mumol modulates the action of SP on the activities of the investigated enzymes.
Gen Pharmacol 1985
PMID:Role of theophylline during the action of SP in vitro on the activity of synaptosomal membrane ATPase (EC 3.6.1.3) from different areas of rat brain. 258 49


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