The f1 portion of H+-translocating ATPase was purified from membrane vesicles of Vibrio parahaemolyticus by a rapid procedure. The whole purification process (from culture of cells to purification of the enzyme) could...
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The f1 portion of H+-translocating ATPase was purified from membrane vesicles of Vibrio parahaemolyticus by a rapid procedure. The whole purification process (from culture of cells to purification of the enzyme) could be completed in 1 day. The f1-ATPase consists offive subunits (.alpha., .beta., .gamma., .delta. and .epsilon.) like f1 of Escherichia coli and other microorganisms. The f1-ATPase of V. parahaemolyticus showed some interesting properties. Its activity was greatly stimulated by high concentrations (about 0.5 M) of SO42-, SO32- and CH3COO-, their effects decreasing in this order. Among the anions tested, Cl- and NO3- were ineffective, or rather inhibitory, and cations had no significant effects. Ethanol (or methanol) stimulated the activity 2- to 3-fold. The activity was inhibited by 4-acetamido-4''-isothiocyanostilbene 2,2''-disulfonate (SITS) (an anion exchanger inhibitor), tetrachlorosalicylanilide (TCS) (an H+ conductor), azide and N-ethylmaleimide. Zinc inhibited the activity only slightly, although it strongly inhibited the ATPase activity in membrane vesicles.
It is shown that ATP dissociates very slowly ( k off 3 h) from the three noncatalytic sites of E. coli f 1 -ATPase and that ADP dissociates from these three sites in a homogeneous fashion with k off = 1.5 × 10 −...
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It is shown that ATP dissociates very slowly ( k off < 6.4 × 10 5 s −1 , t 1 2 > 3 h) from the three noncatalytic sites of E. coli f 1 -ATPase and that ADP dissociates from these three sites in a homogeneous fashion with k off = 1.5 × 10 −4 s −1 ( t 1 2 = 1.35 h). Mutagenesis of α-subunit residues R171 and Q172 in the ‘glycine-rich loop’ (Homology A) consensus region of the noncatalytic sites was carried out to test the hypothesis that unusually bulky residues at these positions are responsible wholly or partly for the observed tight binding of adenine nucleotides. The mutations αQ172G or αR171S,Q172G had no effects on ATP or ADP binding to or rates of dissociation from f 1 noncatalytic sites. K d ATP and K d ADP of isolated α-subunit were weakened by approximately 1 order of magnitude in both mutants. The results suggest that neither residue αR171 nor αQ172 interacts directly with bound nucleotide, and show that the presence of bulky residues per se in the glycine-rich loop region off 1 -α-subunit is not responsible for tight binding in the noncatalytic sites.
The intracellular transport of newly synthesized beta-subunits of the f1-ATPase (beta f1) and of newly synthesized ADP/ATP carrier was followed in isolated rat hepatoma cells. As tested by rapid fractionation of [35S]...
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The intracellular transport of newly synthesized beta-subunits of the f1-ATPase (beta f1) and of newly synthesized ADP/ATP carrier was followed in isolated rat hepatoma cells. As tested by rapid fractionation of [35S]methionine pulse- and pulse-chase-labeled cells and by sensitivity of labeled polypeptides to externally added protease, the import of beta f1 into mitochondria was strongly inhibited by the additional low concentrations of rhodamine 6G (R6G). In contrast, the import of the ADP/ATP carrier into mitochondria was not affected by the inhibitor. The results imply that the proteolytic processing of the precursor of beta f1 is coupled to its translocation across the mitochondrial membrane.
Nucleotide-depleted mitochondrial f1-ATPase binds 3'-(2')-O-(2-nitro-4-azidobenzoyl)-derivatives of ATP (NAB-ATP) and GTP (NAB-GTP) when these nucleotide analogues are added to the enzyme in equimolar quantiti...
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Nucleotide-depleted mitochondrial f1-ATPase binds 3'-(2')-O-(2-nitro-4-azidobenzoyl)-derivatives of ATP (NAB-ATP) and GTP (NAB-GTP) when these nucleotide analogues are added to the enzyme in equimolar quantities in the presence of Mg2+ (uni-site catalysis conditions). The binding of NAB-ATP is accompanied by its hydrolysis and inorganic phosphate dissociation from the enzyme; NAB-ADP remains bound to f1-ATPase. The f1-ATPase X NAB-ADP complex has no ATPase activity and its reactivation in the presence of an excess of ATP is accompanied by NAB-ADP release. The illumination of the f1-ATPase complexes with NAB-ADP or NAB-GDP leads to the covalent binding of one nucleotide analogue molecule to the enzyme and to the irreversible inactivation off1-ATPase. It follows from the results obtained that the modification of just one of the f1-ATPase catalytic sites is sufficient to complete the inhibition of ATPase activity.
Semecarpus anacardium L.f. has been commonly used in various traditional medicines from ancient times. The nuts have been described in Ayurveda medication systems to treat numerous clinical ailments. However, isolatin...
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Semecarpus anacardium L.f. has been commonly used in various traditional medicines from ancient times. The nuts have been described in Ayurveda medication systems to treat numerous clinical ailments. However, isolating phytochemical constituents from nuts remain challenging and exhibits cytotoxic effects on other cells. In this study, we have standardized procedures for isolating phytochemicals from the leaf extract. The ethyl acetate leaf extract selectively affects cancer cells in a dose-dependent manner (IC50: 0.57 mu g/ml in MCf-7 cells) in various cancer cell lines and induces apoptosis in cancer cells. However, the non-malignant cells were relatively insensitive to the ***, the incubation of the leaf extract induces cell cycle arrest and suppresses cancer cell migration in the cell culture model. Moreover, oral administration of extract significantly restored tumor growth in mice. Together, these observations suggest the anti-cancer activities of S. anacardium L.f. leaf potential for both in vitro and in vivo models.
The concentrations of n-butanol and tetracaine required for 50% inhibition of the ATPase activity off1 particles isolated from bovine heart mitochondria were 160 mM and 1.1 mM, respectively. The physiological effects...
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The concentrations of n-butanol and tetracaine required for 50% inhibition of the ATPase activity off1 particles isolated from bovine heart mitochondria were 160 mM and 1.1 mM, respectively. The physiological effects of the anesthetics may be due to direct interaction with membrane proteins rather than with the lipids.
The (n, f, k): f/n, k, f: f system is the combination of most popular consecutive-k-out-of-n and f-out-of-n system and its failure is caused by two different failure criteria. The (n, f, k): f (n, k, f: f) system cons...
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The (n, f, k): f/n, k, f: f system is the combination of most popular consecutive-k-out-of-n and f-out-of-n system and its failure is caused by two different failure criteria. The (n, f, k): f (n, k, f: f) system consists of n components ordered in a line or circle, while the system fails if and only if there exist at least ffailed components or (and) at least k consecutive failed components. In this paper, we consider the sequence {X-u, u 1} of {0, 1}-valued Markov Bernoulli trials (MBT) and study the Birnbaum reliability importance for components of (n, f, k): f(G), and n, k, f: f(G) system through the conditional joint distribution off="lsta_a_749285_***"> f="lsta_a_749285_***">, where f="lsta_a_749285_***"> and f="lsta_a_749285_***"> is the number of non overlapping occurrences of i-runs of length k(i)(i = 0, 1) in n MBT. The formula for evaluation of exact Birnbaum reliability importance is developed and the results are demonstrated numerically. We also bring out the important inter-relations between the reliability and reliability importance offour systems as f-out-of-n: f, consecutive-k-out-of-n: f, (n, f, k): f and n, k, f: f systems.
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