Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: 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)

1. The binding of [14C]-dicyclohexylcarbodiimide to membrane proteins of beef heart mitochondria has been investigated using dodecylsulphate/polyacrylamide gel electrophoresis. Upon incubation of submitochondrial particles with low concentrations of dicyclohexylcarbodiimide (5 nmol/mg protein) radioactivity was incorporated into three components with apparent molecular weights of 30000, 18000 and less than 6500. Only the two smaller components were found to be extracted into chloroform/methanol. The same two components were labelled when the isolated ATPase complex or a reconstituted F0F1 system was incubated with low concentrations of dicyclohexylcarbodiimide. High concentrations of dicyclohexylcarbodiimide (20-100 nmol/mg protein) resulted in binding to several mitochondrial proteins. 2. The maximal amount of dicyclohexylcarbodiimide which can bind to submitochondrial particles, the isolated ATPase complex, and the reconstituted F0F1 system was found to exceed the amount required for maximal inhibition of the ATPase activity by several-fold. The distribution of the bound [14C]dicyclohexylcarbodiimide between the different dicyclohexylcarbodiimide-binding components was investigated as a function of dicyclohexylcarbodiimide concentration. The smallest and largest components revealed a high affinity for dicyclohexylcarbodiimide-binding which paralleled the inhibition of ATPase activity. The intermediate component had a markedly lower affinity for dicyclohexylcarbodiimide-binding. 3. The larger dicyclohexylcarbodiimide-binding component of the isolated ATPase complex can be converted into the smaller component by treatment of the ATPase complex with performic acid. Partial conversion can also be achieved by extraction of the band from the dodecylsulphate-polyacrylamide gel after electrophoresis, followed by re-electrophoresis. The observations suggest that the larger component may be an oligomer of the smaller one. 4. Using concentrations of oligomycin and dicyclohexylcarbodiimide which were equal to or greater than those required for maximal inhibition of the ATPase activity, oligomycin was found to diminish the binding of [14C]dicyclohexylcarbodiimide to both dicyclohexylcarbodiimide-binding components of the isolated ATPase complex.
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PMID:A study of the dicyclohexylcarbodiimide-binding component of the mitochondrial ATPase complex from beef heart. 645 2

Use of the Flagyl selection procedure [Schmidt et al. (1977) Proc. Natl Acad. Sci. USA, 74, 610-614] led to the isolation of a nuclear mutant of Chlamydomonas reinhardtii designated thm-24. This mutant displays normal electron transport rates in vitro, possesses high latent ATPase activity bound to the thylakoid membrane, but is incapable of photophosphorylation. Decay of the transmembrane potential, as indicated by the kinetics of the 520-nm absorption change after illumination, is unusually slow and markedly biphasic. Sodium dodecylsulfate/polyacrylamide gel electrophoresis of purified thylakoid membranes shows mutant thm-24 to be lacking a number of polypeptides including those previously designated 4.1, 4.2 and 8.1. Treatment of purified thylakoid membranes of wild-type and mutant algae, using the chloroform-release procedure of Beechey et al. [(1975) Biochem. J. 148, 533-537] resulted in the removal of ATPase activity from each strain. In wild-type cells, the ATPase activity was of heterogeneous enzymatic origin; fractionation of the chloroform-release extracts by non-denaturing polyacrylamide gel electrophoresis yielded three distinct bands displaying ATPase activity, designated ATPases I, II and III. In contrast, extracts from membranes of mutant thm-24 yielded only one ATPase-containing fraction, co-migrating with ATPase I from wild-type. Use of electrophoretic, immunological and enzymatic methods established a correspondence of the polypeptide subunits of ATPases II and III and those of spinach coupling factor, CF1. ATPase I from either algal strain was shown to be structurally distinct from high plant CF1 and to C. reinhardtii ATPases II and III.
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PMID:A nuclear mutant of Chlamydomonas reinhardtii defective in photosynthetic photophosphorylation. Characterization of the algal coupling factor ATPase. 645 94

Chloroplast thylakoid particles were prepared from wild-type Chlamydomonas reinhardi by gentle sonication. These particles catalyzed phenazine methosulfate dependent photophosphorylation with rates ranging from 300 to 700 mumol of adenosine 5'-triphosphate (ATP) formed (mg of chlorophyll)-1h-1. Photophosphorylation was not sensitive to tentoxin but was sensitive to an anticoupling factor 1 (CF1) antiserum preparation made against spinach CF1. The C. reinhardi chloroplast CF1 was isolated from thylakoid particles by either chloroform or ethylenediaminetraacetic acid extraction. The former enzyme appeared to be missing the gamma subunit and did not reconstitute with partially resolved thylakoid particles. The latter enzyme reconstituted with partially resolved particles and had a specific activity at 37 degrees C of 2-5 umol of ATP hydrolyzed (mg of protein)-1 min-1. The enzyme utilized both MnATP and MgATP. CaATP was a poor substrate, and SrATP was not hydrolyzed. The enzyme was not activated by heat or proteolysis but was stimulated approximately 2-fold by 50 mM dithiothreitol. Alcohols reversibly stimulated the ATPase activity of the enzyme 5-25-fold. Ethanol, 20%, dramatically lowered the temperature optimum from approximately 75 to approximately 45 degrees C and slightly lowered the pH optimum from 8.5 to 8.2. Ethanol had no effect on the activation energy of the ATPase reaction (17 +/- 1.7 kcal/mol). The kinetics of the ATPase reaction catalyzed by the C. reinhardi enzyme are complex. Both free divalent cations and divalent cation ATP inhibited the activity of the enzyme. The apparent Km for MgTAP (55 uM free Mg2+) was approximately 0.2 mM.
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PMID:Isolation, purification, and characterization of coupling factor 1 from Chlamydomonas reinhardi. 645 33

1. Preincubation of the ox heart chloroform-released mitochondrial ATPase with MgATP results in a time-dependent inhibition of ATPase activity. No re-activation occurs when MgATP remains in the preincubation medium. The enzyme activity returns when all the MgATP in the preincubation system has been hydrolysed. 2. The mechanism of the MgATP-induced inhibition was examined. Inhibition occurs on incubation with MgATP or other hydrolysable nucleotides. Incubation with MgADP or Pi does not cause any inhibition. Neither freshly bound adenine nucleotide nor Pi is associated with inhibited enzyme. The rate of MgATP-induced inhibition correlates with the rate of ATP hydrolysis in the preincubation medium. Changing the rate of ATP hydrolysis at a fixed concentration of ATP also changes the rate of MgATP-induced inhibition by the same proportion. The inhibition is thus related to the ATP-hydrolysis process itself. 3. We propose that intermediate enzyme species of the ATP-hydrolytic sequence can undergo a conformational change to form inhibited species. The kinetics of the inhibition suggest that a substrate-activation step is involved in ATP hydrolysis and MgATP-induced inhibition. 4. The effects of the nature of the preincubation medium on the process of MgATP-induced inhibition and its reversal were examined.
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PMID:MgATP-induced inhibition of the adenosine triphosphatase activity of the chloroform-released mitochondrial adenosine triphosphatase. 645 83

The membrane-bound ATPase activity of Bacillus subtilis was inhibited by dicyclohexylcarbodiimide (DCCD). The DCCD-reactive proteolipid of B. subtilis was extracted, from labelled or untreated membranes containing F1 or depleted of F1, with neutral or acidic chloroform/methanol. Purification of the [14C]DCCD-binding proteolipid was attempted by column chromatography on methylated Sephadex G-50 and on DEAE-cellulose. The maximal amount of DCCD which could be bound to the purified proteolipid was found to exceed the amount bound by the purified proteolipid extracted from membranes labelled with the lowest [14C]DCCD concentration required for maximal inhibition of the membrane-bound ATPase activity. The radioactive protein peaks eluted by gel filtration and ion-exchange chromatography were analysed by urea-SDS polyacrylamide slab gel electrophoresis and autoradiography. Radioactivity was incorporated into two components of Mr 18 000 and 6000 when proteolipid was purified by methylated Sephadex. The 6000 polypeptide was always present, whatever the extraction and purification procedures. However, the 18 000 polypeptide was present in largest quantity only when proteolipid was extracted from membranes containing F1 and purified by methylated Sephadex. When proteolipid was purified on DEAE-cellulose this [14C]DCCD binding component of Mr 18 000 was absent.
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PMID:Isolation and purification of dicyclohexylcarbodiimide-reactive proteolipid from Bacillus subtilis membrane. 646 55

The general anesthetics chloroform and halothane inhibit ATP synthesis in rat liver mitochondria, in the millimolar concentration range (1-12 mM), in parallel with a reduction of respiratory control and the ratio of ATP produced to oxygen consumed. In these effects, halothane and chloroform are similar to classical, protonophoric, uncouplers. The rate of ADP-stimulated respiration or the rate of uncoupler-stimulated respiration is not affected. Like classical uncouplers, halothane and chloroform also stimulate mitochondrial ATPase activity. However, the extent of stimulation by these agents is larger than by protonophoric uncouplers and, more significantly, ATPase activity stimulated by carbonylcyanide m-chlorophenylhydrazone is further stimulated by these agents. In the presence of the Ca2+ chelator EGTA, halothane and chloroform have no measurable effect on the magnitude of the proton electrochemical potential, delta mu H. In the absence of EGTA these anesthetics have a small effect on delta mu H, apparently due to stimulation of Ca2+ cycling. Under these conditions the membrane potential is decreased while delta pH is increased, but the total value of delta mu H is only slightly decreased. The uncoupling activity of the anesthetics is the same in the presence of absence of EGTA. Thus, in contrast to protonophoric uncouplers, the uncoupling effect of general anesthetics does not depend on the collapse of delta mu H. In the same concentration range in which anesthetics uncouple oxidative phosphorylation both halothane and chloroform increase membrane fluidity, as measured by the partitioning of the hydrophobic spin probe 5-doxyldecane. These findings suggest a role for intramembrane processes in energy conversion that is not dependent on the bulk delta mu H.
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PMID:Uncoupling of oxidative phosphorylation in rat liver mitochondria by general anesthetics. 657 86

African swine fever virus polypeptides with molecular weight of 120, 78, 69, 59, 56, 45, 39, 28, 26, 24, 16, and 14 kD are the major proteins in the purified virions, as shown by electrophoresis and immunoblotting. A mixture of proteases and pancreatic lipase hydrolyzed the polypeptides of 120 and 78 kD in viral preparations at low concentrations of enzymes, polypeptides of 69, 56, 45, 39, 28, and 14 kD disappeared after treatment with this mixture at medium concentrations, and 26 kD polypeptide was eliminated at a high concentration of the enzymes. The 21 kD polypeptide which did not react with the specific antiviral serum in immunoblotting was not hydrolyzed by proteases contaminating lipase. Treatment with triton X-100 and ether boosted the activity of DNA-dependent RNA-polymerase, whereas treatment with ether followed by resedimentation markedly decreased polymerase activity in the resultant sediment. Treatment with diethyl ether did not influence the activity of virus-associated ATPase, which was partially resistant to denaturating organic solvents acetone and chloroform-methanol mixture. Our findings and published data permitted us to propose a schematic arrangement of viral polypeptides and enzymes in the virion structure.
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PMID:[Localizing the major peptides of African swine fever virus and virus-associated enzymes in the virion structure]. 747 38

Membrane fractions and chloroform-methanol (C-M) extracts of jimpy (jp) and normal CNS at 17-20 days were examined by immunoblot and sequence analysis to determine whether myelin proteolipid protein (FLP) or DM-20 could be detected in jp CNS. No reactivity was detected in jp samples with several PLP antibodies (Abs) except with one Ab to amino acids 109-128 of normal PLP. Proteins in the immunoreactive bands approximately 26 M(r) comigrating with PLP were sequenced for the first 10-12 residues. A sequence corresponding to PLP was found in normal CNS, as expected, but not in the band from jp CNS. Our results provide no evidence for an aberrant form of PLP in jp CNS at 17-20 days. This and other studies suggest that the abnormalities in jp brain are not due to toxicity of the mutant jp PLP/DM-20 proteins. Interestingly, a sequence identical to the amino terminus of the mature proton channel subunit 9 of mitochondrial F0 ATPase was detected in the immunoreactive bands approximately 26 M(r) in both normal and jp samples. This identification was supported by reactivity with an Ab to the F0 subunit and by labeling with dicyclohexylcarbodiimide (DCCD). In contrast to PLP isolated from whole CNS, PLP isolated from myelin was devoid of F0 subunit 9 based on sequence analysis and lack of reactivity with an Ab to the F0 subunit, yet still reacted with DCCD. This finding rules out the possibility that contaminating F0 ATPase gives rise to the DCCD binding exhibited by PLP and confirms the possibility that PLP has proton channel activity, as suggested by Lin and Lees (1,2).
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PMID:Analysis of myelin proteolipid protein and F0 ATPase subunit 9 in normal and jimpy CNS. 752 46

Subunit c of the Escherichia coli F1F0-ATPase, purified in chloroform/methanol (2:1), was reconstituted with detergent-solubilized F0 subunits a and b to form a functionally active H+ channel. The rates of H+ uptake by the proteoliposomes containing the reconstituted F0 complex were comparable to those observed with native F0 reconstituted without subunit dissociation. The F0 reconstituted from purified subunits was also shown to form an active ATP-driven H+ pump upon binding of the F1-ATPase sector of the complex. Reconstitution of D61N and D61G mutant c subunits with wild-type subunits a and b produced an inactive F0. Hybrid F0 complexes, formed with mixtures of wild-type and D61N or D61G mutant c subunits, were also prepared. Formation of an active F0 was prevented by addition of relatively small proportions of D61N or D61G mutant c subunits, i.e. active F0 formation was gradually disrupted as the mutant/wild-type ratio was increased from 0.05 to 0.2. The hybrid reconstitution studies support a model where inactivation of one of the 9-12 c subunits found in F0 is sufficient to abolish activity.
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PMID:Reconstitution of the Fo complex of Escherichia coli ATP synthase from isolated subunits. Varying the number of essential carboxylates by co-incorporation of wild-type and mutant subunit c after purification in organic solvent. 758 91

Adriamycin (AD)-Fe3+ caused the inactivation of Na(+)-, K(+)-ATPase and Ca(2+)-ATPase of erythrocyte membranes during lipid peroxidation. AD-Fe3+ also induced the formation of fluorescent substances from the membranes with lipid peroxidation. The fluorescent substances were little extracted by chloroform-methanol, indicating that they were retained in the membranes. Butylated hydroxytoluene and trolox strongly inhibited both the inactivation of these ATPases and the formation of fluorescent substances with lipid peroxidation. Another antioxidant, vitamin E, slightly prevented the damage of the membranes. However, p-nitrophenyl phosphatase activity and acetylcholine esterase have lower or no susceptibility to the membrane lipid peroxidation. These results indicated that the ATPases were very sensitive to lipid peroxidation and that the membranes were modified during the peroxidation reaction.
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PMID:Adriamycin-Fe(3+)-induced inactivation of enzymes in erythrocyte membranes during lipid peroxidation. 774 51


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