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Biomedical subjects

Y Hatefi

Publications and source records attributed to Y Hatefi.

At least 127 records · Page 7Linked to original sources
PubMed1977

Dehydrogenase and transhydrogenase properties of the soluble NADH dehydrogenase of bovine heart mitochondria.

The soluble NADH dehydrogenase of low molecular weight, isolated from complex I (NADH:ubiquinone oxidoreductase, EC 1.6.5.3) of the respiratory chain, has been shown to have NADPH dehydrogenase and NADPH leads to NAD transhydrogenase activities. Both activities are greatly increased in the presence of added guanidine-HCl and at pH values less than 6.5. The chromophores of the soluble enzyme (flavin and iron--sulfur centers) are reduced by NADH and NADPH to the same extent. The latter reduction is extremely slow, and is considerably stimulated in the presence of guanidine-HCl. The soluble dehydrogenase has little or no NADH leads to NADP and NADPH leads to NADP transhydrogenase activity. The former reaction is known to be energy-linked in submitochondrial particles; the latter was shown in the present studies also to be energy-linked. In view of the above and earlier results, possible mechanisms for dehydrogenation and transhydrogenation (nonenergy-linked and energy-linked) involving reduced and oxidized NAD and NADP are proposed.

Animals↗
PubMed1976

Oxidative phosphorylation properties of mitochondria isolated from transplanted hepatoma.

Mitochondria were isolated from Morris hepatomas with rapid (types 3683, 7777, and 3924A) and intermediate (types 5123D and 7800) growth rates, using proteolytic digestion of minced tumor tissue to release the particles. Mitochondria isolated by the same procedure from rat liver were employed as controls. All the hepatoma mitochondria were capable of coupled respiration with normal phosphorylation yields (ADP/O) and respiratory control ratios ranging from 2 to considerably more than 10. Particles from hepatomas 7777 and 7800 exhibited properties closest to liver mitochondria, while those from hepatomas 3683 and 3924A showed the greatest difference. All the hepatoma mitochondria were capable of oxidizing succinate, 3-hydroxybutyrate and monoamines. However, the oxidation rates of the latter two substrates by mitochondria from hepatomas 3683 and 3924A were only a fraction of the control rates. These differences appeared to be due, at least in part, to the structural instability of the isolated hepatoma mitochondria. In contrast to the reports of others, all hepatoma mitochondria exhibited considerable stimulation of ATPase activity by uncouplers. Maximal stimulation of ATPase activity by representatives of three classes of uncouplers was in all instances comparable to the values obtained for rat liver mitochondria.

Acid Phosphatase↗
PubMed1976

Purification and properties of a low molecular weight protein factor of mitochondrial energy-linked functions.

1. A soluble protein with a molecular weight of 11-12-10(3) has been isolated from bovine-heart mitochondria, which stimulates the following ATP-dependent reactions of submitochondrial particles treated with 0.6 mM EDTA and 1 M NH4OH: reverse electron transfer from succinate to NAD, transhydrogenation from NADH to NADP, and ATP-Pi exchange. The factor has no effect on the NADH oxidase, succinate oxidase and ATPase activities of the particles. 2. The stimulatory effect of the factor in the ATP-dependent reduction of NAD by succinate is 12 mumol-min-1-mg-1 of the factor protein. However, the NH4OH-EDTA treated particles are saturated for maximal activation of the above reaction by very small amounts of the factor (about 20-40 mug factor per mg particle). 3. Electrophoresis of the factor preparation on polyacrylamide gels showed a single protein band plus a nonprotein material which moved at the dye front and was weakly stained with Coomassie Blue. The protein was shown to be required for activation of the particles; whether the fast-moving, nonprotein material is also required is not known. 4. The factor is inhibited by mercurials and N-ethylmaleimide. The former, but not the latter, inhibition is completely reversed by 1,4-dithiothreitol. 5. The NH4OH-EDTA treated particles are also stimulated by rutamycin up to about 0.1 nmol of rutamycin per mg particle; higher rutamycin concentrations inhibit. Depending on the particle preparation, the factor stimulates up to about 3 nmol per mg particle, but does not inhibit at higher concentrations. In addition, under certain conditions in which appropriate concentrations of rutamycin fail to stimulate the particles, the factor still does.

4-Chloromercuribenzenesulfonate↗
PubMed1975

Energy conservation and uncoupling in mitochondria.

Energy conservation and uncoupling in mitochondria are examined in the light of three important new findings: (a) Studies with the photoaffinity-labeling uncoupler 2-azido-4-nitrophenol have shown that mitochondria contain a specific uncoupler binding site (apparently a polypeptide of Mr = 30,000 +/- 10%). (b) This site fractionates into an enzyme complex (complex V), which is capable of oligomycin- and uncoupler-sensitive ATP-Pi exchange. It is absent from electron transfer complexes I, III, and IV, which represent segments of the respiratory chain containing coupling sites 1, 2, and 3, respectively. (c) Trinitrophenol is a membrane-impermeable uncoupler (uncouples submitochondrial particles, but not mitochondria) and a poor protonophore. There is an excellent correlation between the uncoupling potencies and the affinities of uncouplers for the mitochondrial uncoupler-binding site. There is no correlation between uncoupling potency and protonophoric activity of uncouplers when a membrane-permeable uncoupler is compared with a membrane-impermeable one.

Adenosine Triphosphate↗
PubMed1975

Mitochondrial ATP-Pi exchange complex and the site of uncoupling of oxidative phosphorylation.

Five enzyme complexes, which are concerned with electron transport and oxidative phosphorylation, have been isolated from beef heart mitochondria. Enzyme complexes I, II, III and IV are the electron transfer complexes discovered in 1961. Complex V is an energy-conserving complex. It catalyzes ATP-Pi exchange and ATP hydrolysis. The exchange reaction is sensitive to uncouplers, rutamycin, valinomycin plus K-+, dicyclorexylcarboditmide, arsenate, azide, and adenylyl imidodiphosphate. It is also specific for ATP; ITP, GTP and UTP are essentially ineffective. Studies with the photoaffinity labeling uncoupler, 2-azido-4-nitrophenol (NPA), have shown that the mitochondrial uncoupler-binding sites are located exclusively in complex V. Complexes I, III and IV, which carry the three coupling sites of the respiratory chain, had negligible capacity for the binding of NPA, whereas the uncoupler-binding capacity of complex V appeared to be increased two- to threefold as compared to mitochondria. Complexes I, II, III, IV and V are obtained from the same batch of mitochondria by a simple fractionation procedure, which employs cholate, deoxycholate, ammonium acetate and ammonium sulfate. Studies with NPA have shown that mitochondria contain per milligram protein about 0.6 nmole of uniformly reacting uncoupler binding site. All of the uncouplers tested appeared to interact competitively with this site. Photoaffinity labeling with tritiated NPA has shown that a major portion of NPA binds to a polypeptide of molecular weight between 26,000 and 30,000. Other studies on the mechanism of uncoupling have shown that picrate is a membrane-impermeable uncoupler. It cannot uncouple mitochondria. However, it is an effective uncoupler of ATP synthesis and ATP-induced transhydrogenation or reverse electron transfer when used in conjunction with sonicated submitochondrial particles, which have an inside-out orientation of the inner membrane with respect to the medium. In these particles, picrate binds to the same uncoupler-binding site as NPA and other uncouplers. However, unlike the membrane-permeable uncouplers, picrate is a poor protonophore. It has a very small effect on the proton permeability of phosphorylating submitochondrial vesicles, even at two to three times the concentration needed for complete uncoupling. The increase in the proton permeability of submitochondrial vesicles caused by such high concentrations of picrate (500 mum) can be achieved with approximately 5 mum 2,4-dinitrophenol. At this concentration, dinitrophenol results in only about 20% uncoupling.

Adenosine Triphosphate↗