共查询到20条相似文献,搜索用时 15 毫秒
1.
L A Novikova S M Dracheva A S Zubatov V N Luzikov 《Biokhimii?a (Moscow, Russia)》1982,47(8):1401-1408
The proteins of submitochondrial particles solubilized with 0.1% Triton X-100 were separated by polyacrylamide gel electrophoresis. Hydrolysis of several proteinase substrates was registered directly in the gel after completion of electrophoresis. According to the data obtained the inner mitochondrial membrane contains one or two enzymes which catalyze hydrolysis of cytochrome c as well as one or two enzymes splitting synthetic substrate of trypsin-like proteinases, e. g. N-alpha-benzoyl-L-arginine-p-nitroanilide (BAPA) and N-alpha-benzoyl-L-arginine-beta-naphthylamide (BANA). Submitochondrial particles were shown to catalyze hydrolysis of 3H-labelled cytochrome c. This activity is suppressed by the same inhibitors as the hydrolysis of mitochondrial translation products, i. e. phenyl-methylsulfonylfluoride, p-chloromercuribenzosulfonate, leupeptin and antipain. Presumably these two processes are catalyzed by the same enzyme localized in the inner mitochondrial membrane. Physiological functions of BAPA- and BANA-hydrolyzing enzyme(s) are still unclear. 相似文献
2.
Cruciat CM Brunner S Baumann F Neupert W Stuart RA 《The Journal of biological chemistry》2000,275(24):18093-18098
The mitochondrial electron transport chain complexes are large multisubunit complexes embedded in the inner membrane. We report here that in the yeast Saccharomyces cerevisiae, the cytochrome bc(1) and cytochrome c oxidase complexes co-exist as a larger complex of approximately 1000 kDa in the mitochondrial membrane. Following solubilization with a mild detergent, the cytochrome bc(1)-cytochrome c oxidase complex remains stable. It was analyzed using the techniques of gel filtration and blue native-polyacrylamide gel electrophoresis. Direct physical association of subunits of the cytochrome bc(1) complex with those of the cytochrome c oxidase complex was verified by co-immunoprecipitation analysis. Our data indicate that the cytochrome bc(1) complex is exclusively in association with the cytochrome c oxidase complex in yeast mitochondria. We term this complex the cytochrome bc(1)-cytochrome c oxidase supracomplex. 相似文献
3.
The aorta in male mice shows higher activities of several lysosomal hydrolases and of cytochrome c oxidase, an inner mitochondrial membrane enzyme, than in female mice. Orchiectomy abolishes this sex difference, whereas testosterone administration induces an accretion of RNA and protein and elevated activities of lysosomal hydrolases and cytochrome c oxidase. However, the outer mitochondrial membrane enzyme monoamine oxidase is unaffected by sex, orchiectomy or testosterone. Thus, androgens regulate cell growth and enzymes associated with lysosomes and the inner mitochondrial membrane. 相似文献
4.
Cytochrome c release from mitochondria is a critical event in the apoptosis induction. Dissociation of cytochrome c from the mitochondrial inner membrane (IMM) is a necessary first step for cytochrome c release. In the present study, the effect of reactive oxygen species (ROS) on the dissociation of cytochrome c from beef-heart submitochondrial particles (SMP) and on the cardiolipin content was investigated. Exposure of SMP to mitochondrial-mediated ROS generation resulted in a large dissociation of cytochrome c from SMP and in a parallel loss of cardiolipin. Both these effects were directly and significantly correlated and also abolished by superoxide dismutase+catalase. These results demonstrate that ROS generation induces the dissociation of cytochrome c from IMM via cardiolipin peroxidation. The data may prove useful in clarifying the molecular mechanism underlying the release of cytochrome c from the mitochondria to the cytosol. 相似文献
5.
1. Mitochondria were isolated from developing endosperm of Ricinus communis and were fractionated into outer membrane and inner membrane. The relative purity of the two membrane fractions was determined by marker enzymes. The fractions were also examined by negative-stain electron microscopy. 2. Membrane fractions were sequentially extracted in the following way. (a) Suspension in 0.5M-potassium phosphate, pH7.1; (b)suspension in 0.1M-EDTA (disodium salt)/0.05M-potassium phosphate, pH7.1; (c) sonication in 0.05M-potassium phosphate, pH7.1;(d)sonication in aq. Triton X-100 (0.1%). The membranes were pelleted by centrifugation at 100 000g for 15 min, between each step. Agglutination activity in the extracts was investigated by using trypsin-treated rabbit erythrocytes. 3. The addition of lactose to inner mitochondrial membrane resulted in the solubilization of part of the lectin activity, indicating that the protein was attached to the membrane via its carbohydrate-binding site. Pretreatment of the membranes with lactose before tha usual extraction procedure showed that lactose could extract lectins that normally required more harsh treatment of the membrane for solubilization. 4. Lectins extracted from inner membranes were purified by affinity chromatography on agarose gel. Polyacrylamide-gel electrophoresis of purified samples in sodium dodecyl sulphate indicated that at least part of the lectin present in inner mitochondrial membrane was identical with the R. communis agglutinin of mol.wt. 120 000. 相似文献
6.
Correlated three-dimensional light and electron microscopy reveals transformation of mitochondria during apoptosis 总被引:4,自引:0,他引:4
Sun MG Williams J Munoz-Pinedo C Perkins GA Brown JM Ellisman MH Green DR Frey TG 《Nature cell biology》2007,9(9):1057-1065
In addition to their role in cellular bioenergetics, mitochondria also initiate common forms of programmed cell death (apoptosis) through the release of proteins such as cytochrome c from the intermembrane and intracristal spaces. The release of these proteins is studied in populations of cells by western blotting mitochondrial and cytoplasmic fractions of cellular extracts, and in single cells by fluorescence microscopy using fluorescent indicators and fusion proteins. However, studying the changes in ultrastructure associated with release of proteins requires the higher resolution provided by transmission electron microscopy. Here, we have used fluorescence microscopy to characterize the state of apoptosis in HeLa cells treated with etoposide followed by electron microscopy and three-dimensional electron microscope tomography of the identical cells to study the sequence of structural changes. We have identified a remodelling of the inner mitochondrial membrane into many separate vesicular matrix compartments that accompanies release of proteins; however, this remodelling is not required for efficient release of cytochrome c. Swelling occurs only late in apoptosis after release of cytochrome c and loss of the mitochondrial membrane potential. 相似文献
7.
F di Jeso S Bartolucci D Giorgini 《Comptes rendus des séances de la Société de biologie et de ses filiales》1978,172(2):274-277
A specific antibody against cytochrome c1 (pig heart mitochondria) has been obtained. It inhibits the electron transport of the respiratory chain in the intact mitochondria at the cytochrome c1 site of inner mitochondrial membrane ; but it has no effect on the isolated submitochondrial particles (inside-out inner mitochondrial membrane vesicles free of any outer membrane or outside-out inner membrane). Thus the topologic position of cytochrome c1 in the inner mitochondrial membrane is asymetrically lcoated on the outer side of the inner mitochondrial membrane. These results agree with our previous researches on ATP-ase and cytochromes b, c and a, indicating the location on the inner side for the first one, transmembranous for the last one, on the outer side for the others respiratory chain components. Thus the electron transport from cytochrome b to a takes place in the outer region of inner mitochondrial membrane and the transmembranous location of cytochrome-oxidase facilitates the transfer of the electrons to oxygen. 相似文献
8.
A K Grover C Y Kwan J Crankshaw E E Daniel 《Canadian journal of physiology and pharmacology》1981,59(11):1128-1133
Isopycnic centrifugation experiments using sucrose density gradients showed that in digitonin-treated microsomes the distribution of the plasma membrane (PM) marker 5'-nucleotidase was shifted to higher densities. The treatment also caused similar but less pronounced changes in the distribution of protein, the putative endoplasmic reticulum (ER) marker NADPH-dependent cytochrome c reductase, and the inner mitochondrial marker cytochrome c oxidase. Similar experiments using more purified membrane fractions showed that the digitonin treatment led to a comparable increase in the densities of the fractions N1 and N2 previously described as subfractions of plasma membrane and to considerably less increase in the density of the fraction N3B which is enriched in the endoplasmic reticulum and the inner mitochondrial markers. Digitonin inhibited the ATP-dependent Ca uptake by the N1 fraction in a concentration-dependent manner (I50 = 0.3 mg/mL). Digitonin (0.5 mg/mL) inhibited the ATP-dependent azide-insensitive Ca uptake by all the fractions. The results support the hypothesis that (a) N1 and N2 are subfractions of plasma membrane, and (b) ATP-dependent azide-insensitive Ca uptake in rat myometrium is a property of plasma membranes. 相似文献
9.
The pro-apoptotic proteins, Bid and Bax, cause a limited permeabilization of the mitochondrial outer membrane that is enhanced by cytosol
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Kluck RM Esposti MD Perkins G Renken C Kuwana T Bossy-Wetzel E Goldberg M Allen T Barber MJ Green DR Newmeyer DD 《The Journal of cell biology》1999,147(4):809-822
During apoptosis, an important pathway leading to caspase activation involves the release of cytochrome c from the intermembrane space of mitochondria. Using a cell-free system based on Xenopus egg extracts, we examined changes in the outer mitochondrial membrane accompanying cytochrome c efflux. The pro-apoptotic proteins, Bid and Bax, as well as factors present in Xenopus egg cytosol, each induced cytochrome c release when incubated with isolated mitochondria. These factors caused a permeabilization of the outer membrane that allowed the corelease of multiple intermembrane space proteins: cytochrome c, adenylate kinase and sulfite oxidase. The efflux process is thus nonspecific. None of the cytochrome c-releasing factors caused detectable mitochondrial swelling, arguing that matrix swelling is not required for outer membrane permeability in this system. Bid and Bax caused complete release of cytochrome c but only a limited permeabilization of the outer membrane, as measured by the accessibility of inner membrane-associated respiratory complexes III and IV to exogenously added cytochrome c. However, outer membrane permeability was strikingly increased by a macromolecular cytosolic factor, termed PEF (permeability enhancing factor). We hypothesize that PEF activity could help determine whether cells can recover from mitochondrial cytochrome c release. 相似文献
10.
Androgens regulate mitochondrial cytochrome c oxidase and lysosomal hydrolases in mouse skeletal muscle. 总被引:1,自引:0,他引:1
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The gastrocnemius, a fast-twitch white muscle, and the soleus, a slow-twitch red muscle, were studied in A/J mice. The specific activities of the lysosomal hydrolases, beta-D-glucuronidase, hexosaminidase, beta-D-galactosidase and arylsulphatase, the inner-mitochondrial-membrane enzyme cytochrome c oxidase, and the outer-mitochondrial-membrane enzyme monoamine oxidase, were greater in the soleus than in the gastrocnemius. The specific activities of the lysosomal hydrolases and cytochrome c oxidase in the gastrocnemius and soleus were substantially higher in male mice than in female mice. Orchiectomy abolished this sex difference. Testosterone increased the activities of the lysosomal hydrolases and cytochrome c oxidase and coincidentally induced muscle hypertrophy and an accretion of protein and RNA, but total DNA remained constant. Monoamine oxidase was unaffected by sex, orchiectomy and testosterone. These findings indicate that endogenous androgens regulate the activity of enzymes associated with lysosomes and the inner mitochondrial membrane, as well as muscle fibre growth in mouse skeletal muscle. 相似文献
11.
Cytochrome c1 is a subunit of ubiquinol--cytochrome c reductase (EC 1.10.2.2). In Neurospora crassa wild type 74A grown in the presence of chloramphenicol, the subunit is inserted only into the bilayer of the mitochondrial inner membranes without associating with other proteins. From these modified membranes a monodisperse (cytochrome c1)-Triton complex was isolated by subjecting the Triton-solubilized membranes to affinity chromatography on immobilized cytochrome c. A water-soluble pentamer of cytochrome c1 was prepared from the (cytochrome c1)-Triton complex by removing the detergent. By limited proteolytic digestion of the cytochrome c1-Triton complex with chymotrypsin, a water-soluble monomeric cytochrome c1 was prepared which has a molecular weight of only 24 000 as compared to 31 000 of the membrane-bound cytochrome c1. The 24 000-Mr cytochrome c1 and the 31 000-Mr cytochrome c1 have same light absorption spectra and cytochrome-c-binding properties. These results are used to propose the following model. Cytochrome c1 consists of a large hydrophilic part and a small hydrophobic part. The hydrophilic part extends from the mitochondrial inner membrane into the intermembrane space. This part carries the heme and interacts with cytochrome c. The hydrophobic part anchors the cytochrome c1 to the bilayer. 相似文献
12.
A carboxyl-terminal hydrophobic region of yeast cytochrome c1 is necessary for functional assembly into complex III of the respiratory chain 总被引:1,自引:0,他引:1
Cytochrome c1 is an amphiphilic protein which binds to the mitochondrial inner membrane, presumably through a hydrophobic region near the carboxyl (C)-terminus. In the preceding study (Hase, T., et al. (1987) J. Biochem. 102, 401-410), two cytochrome c1 mutations were constructed: delta 1 and delta 2 cytochromes c1, in which the C-terminal segments of 17 and 71 residues were replaced by foreign sequences of 20 and 15 residues, respectively. delta 2 cytochrome c1 had lost the putative membrane anchor. The two cytochrome c1 mutants were localized in mitochondria, but succinate-cytochrome c1 reductase activity was detected only in the mitochondria containing delta 1 cytochrome c1. The membrane association of the two mutant molecules as well as that of authentic cytochrome c1 was investigated. These three molecules were firmly attached to mitochondrial membranes and not solubilized on either sonication or sodium carbonate (pH 11) treatment. However, when the membranes were solubilized with Triton X-100, both the delta 1 and authentic cytochromes c1 were extracted from the membranes more easily than delta 2 cytochrome c1. By fractionating cholate extracts of mitochondrial membranes with ammonium sulfate, delta 1 cytochrome c1 was cofractionated with the enzymatic activity of complex III, but delta 2 cytochrome c1 was clearly separated from the complex III fraction. Trypsin treatment of mitochondria and mitoplasts showed that delta 2 cytochrome c1 was exposed to the intermembrane space, with such a topology that its trypsin susceptibility became much higher than that of the authentic molecule.(ABSTRACT TRUNCATED AT 250 WORDS) 相似文献
13.
A membrane-associated NADH dehydrogenase from beef neutrophils was purified to homogeneity, using detergent (cholate plus Triton X-100) extraction and chromatography on DEAE-Sepharose CL-6B, agarose-hexane-NAD, and hydroxylapatite. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed an apparent subunit molecular weight of 17,500, but the enzyme was highly aggregated (Mr greater than 450,000) in nondenaturing gels containing 0.1% Triton X-100. The protein band in nondenaturing gels was also stained for activity using NADH and nitro blue tetrazolium. The enzyme showed greatest electron acceptor activity with ferricyanide (100%), followed by cytochrome c (3.5%), dichloroindophenol (2.7%), and cytochrome b5 (0.34%). No activity was seen with oxygen. The Km values for NADH and ferricyanide were 18 and 9.5 microM, respectively, and NAD+ was a weak competitive inhibitor (Ki = 118 microM). No activity was seen with NADPH. No effects were seen with mitochondrial respiratory inhibitors such as azide, cyanide, or rotenone, but p-chloromercuribenzoate was strongly inhibitory and N-ethylmaleimide was weakly inhibitory. No free flavin was detectable in enzyme preparations. Based upon kinetic, physical, and inhibition properties, this NADH dehydrogenase differs from those previously described in microsomes and erythrocyte plasma membrane. 相似文献
14.
Bax and heart mitochondria: uncoupling and inhibition of respiration without permeability transition
Appaix F Guerrero K Rampal D Izikki M Kaambre T Sikk P Brdiczka D Riva-Lavieille C Olivares J Longuet M Antonsson B Saks VA 《Biochimica et biophysica acta》2002,1556(2-3):155-167
The effects of Bax (full-length, FL, and C-terminal truncated, DeltaC) on respiration rate, membrane potential, MgATPase activity and kinetics of regulation of respiration were studied in isolated rat heart mitochondria and permeabilized cardiomyocytes. The results showed that while both Bax-FL and Bax-DeltaC permeabilized the outer mitochondrial membrane, released cytochrome c and reduced the respiration rate, the latter could be fully restored by exogenous cytochrome c only in the case of Bax-DeltaC, but not in presence of Bax-FL. In addition, Bax-FL but not Bax-DeltaC increased the MgATPase activity, and their effects on the mitochondrial membrane potential were quantitatively different. None of these effects was sensitive to cyclosporin A (CsA).It is concluded that Bax-FL affects both the outer and the inner mitochondrial membranes by: (1) opening large pores in the outer membrane; (2) inhibiting some segments of the respiratory chain in the inner membrane; and (3) uncoupling the inner mitochondrial membrane by increasing proton leak without opening the permeability transition pore (PTP). 相似文献
15.
THE OXIDATION OF EXOGENOUS AND ENDOGENOUS CYTOCHROME C IN MITOCHONDRIA : A Biochemical and Ultrastructural Study 总被引:2,自引:1,他引:1
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The effect of the nonionic detergent Lubrol on the oxidation of endogenous and exogenous cytochrome c by cytochrome oxidase in intact and fragmented mitochondria was studied. Mitochondria and mitochondrial fragments from liver, kidney, heart, and skeletal muscle have been used. Negatively stained preparations of intact mitochondria showed the particles of Fernández-Morán on the matrix side of their inner membrane system: under these conditions, the oxidation rate of externally added cytochrome c was very high, and it was stimulated very poorly by Lubrol. Mechanical fragmentation of liver mitochondria yielded vesicles with a smooth external profile: also under these conditions, the oxidation of externally added cytochrome c was very high, and poorly stimulated by Lubrol. The oxidation of endogenous cytochrome c was also unaffected by Lubrol. On the other hand, fragmentation of heart and skeletal muscle mitochondria yielded vesicles having numerous particles of Fernández-Morán on their external profiles. Under these conditions, the oxidation of exogenous cytochrome c was low and was markedly stimulated by Lubrol. On the contrary, no activation of the oxidation of endogenous cytochrome c was induced by the detergent. The results indicate a difference in the permeability properties of the two faces of the inner mitochondrial membrane: a permeability barrier for cytochrome c is suggested to exist at the inner face. 相似文献
16.
The role of a mitochondrial pathway in the induction of apoptosis by chemicals extracted from diesel exhaust particles 总被引:10,自引:0,他引:10
Hiura TS Li N Kaplan R Horwitz M Seagrave JC Nel AE 《Journal of immunology (Baltimore, Md. : 1950)》2000,165(5):2703-2711
We are interested in the cytotoxic and proinflammatory effects of particulate pollutants in the respiratory tract. We demonstrate that methanol extracts made from diesel exhaust particles (DEP) induce apoptosis and reactive oxygen species (ROS) in pulmonary alveolar macrophages and RAW 264.7 cells. The toxicity of these organic extracts mimics the cytotoxicity of the intact particles and could be suppressed by the synthetic sulfhydryl compounds, N-acetylcysteine and bucillamine. Because DEP-induced apoptosis follows cytochrome c release, we studied the effect of DEP chemicals on mitochondrially regulated death mechanisms. Crude DEP extracts induced ROS production and perturbed mitochondrial function before and at the onset of apoptosis. This mitochondrial perturbation follows an orderly sequence of events, which commence with a change in mitochondrial membrane potential, followed by cytochrome c release, development of membrane asymmetry (annexin V staining), and propidium iodide uptake. Structural damage to the mitochondrial inner membrane, evidenced by a decrease in cardiolipin mass, leads to O-*2 generation and uncoupling of oxidative phosphorylation (decreased intracellular ATP levels). N-acetylcysteine reversed these mitochondrial effects and ROS production. Overexpression of the mitochondrial apoptosis regulator, Bcl-2, delayed but did not suppress apoptosis. Taken together, these results suggest that DEP chemicals induce apoptosis in macrophages via a toxic effect on mitochondria. 相似文献
17.
Integral membrane proteins from human red cells were fractionated in sodium dodecyl sulfate solutions by high performance gel filtration on the small-bead cross-linked agarose gel Superose 6. The components were identified by acrylamide gel electrophoresis in the presence of sodium dodecyl sulfate. The combination of Superose chromatography with electrophoresis afforded high resolution. As expected the gel filtration elution volumes depended essentially on the molecular mass, but the elution volumes decreased stepwise as the detergent concentration was increased from 0.6 to 100 mM, with the largest decrease for the glucose transporter. The resolution increased as the flow rate was decreased from 60 to 1 ml X cm-2 X h-1. The Mr values for the anion and glucose transporters as estimated by Superose 6-chromatography at 50 mM detergent were 75-80% of the corresponding Mr values obtained by electrophoresis. At 50 mM dodecyl sulfate the proteins were resolved into four fractions (a-d) which mainly contained: (a) dimer and (b) monomer of the anion transporter, (c) the glucose transporter and (d) components of Mr below 40 000. Monoclonal antibodies that possibly are directed against the glucose transporter (Lundahl, P., Greijer, E., Cardell, S., Mascher, E. and Andersson, L. (1986) Biochim. Biophys. Acta 855, 345-356) interacted only with part of the 4.5-material in fraction c in immunoblotting (Western blotting). Superose 6-chromatography of red cell glucose transporter that had been partially purified on DEAE-cellulose and Mono Q resolved one major and two minor fractions. Electrophoretic analysis showed that components of Mr 90,000, 50,000, and 25,000 had been separated from the major Mr-55,000-4.5-material and revealed size heterogeneity within the major chromatographic fraction. Heating of the glucose transporter in the presence of dodecyl sulfate caused an unexpected retardation of monomeric transporter on Superose 6. The apparent Mr decreased from 44,000 to 29,000. 相似文献
18.
Tellis C Pantazi D Ioachim E Galani V Lekka ME 《European journal of cell biology》2003,82(11):573-578
Tetrahymena pyriformis contains platelet-activating factor (PAF) as a minor lipid, which is biosynthesized de novo. A dithiothreitol-insensitive CDP-choline:cholinephosphotransferase (AAG-CPT), which utilizes alkyl-acetyl-glycerol as a substrate, had been detected in both the mitochondrial and microsomal fractions of the protozoan. In the present report, localization of this enzyme in submitochondrial fractions was studied. Cell fractionation was evaluated with enzyme and morphological markers. In this respect, succinate dehydrogenase, NADPH:cytochrome c reductase, glucose-6-phosphatase, alkaline phosphatase, monoaminoxidase, and cytochrome c oxidase activities were investigated. In the presence of antimycin A, mitochondrial activity of NADPH-cytochrome c reductase, was increased, while the microsomal one was reduced. Cardiolipin was distributed in the inner mitochondrial membrane. Alkaline phosphatase was found exclusively in the cytosol of the protozoan. The main portion of the dithiothreitol-insensitive AAG-CPT was localized in the inner mitochondrial membrane. Our data indicate that mitochondria are able to produce PAF, which might be associated with their function. 相似文献
19.
Yeast mitochondria and purified yeast cytochrome c oxidase incorporated into micelles of the nonionic detergent Tween 80 were equilibrated with the hydrophobic aryl azides 5-[125I]iodonaphthyl-1-azide or S-(4-azido-2-nitrophenyl)-[35S]thiophenol. The azides were then converted to highly reactive nitrenes by flash photolysis or by illumination for 2 min and the derivatized cytochrome c oxidase subunits were identified by gel electrophoresis and radioactivity measurements. 5-[125I]Iodonaphthyl-1-azide labeled mainly the three mitochondrially made Subunits I to III and the cytoplasmically made Subunit VII. Subunits IV to VI or cytochrome c bound to the purified enzyme were labeled 9- to 90-fold less. Essentially the same result was obtained with S-(4-azido-2-nitrophenyl)-[35S]thiophenol except that Subunit V was labeled as well. In contrast, all seven subunits as well as cytochrome c were heavily labeled when the enzyme was dissociated with dodecyl sulfate prior to photolabeling with either of the two probes. These data indicate that all subunits of yeast cytochrome c oxidase except Subunits IV and VI are at least partly embedded in the lipid bilayer of the mitochondrial inner membrane. 相似文献
20.
R A Demel W Jordi H Lambrechts H van Damme R Hovius B de Kruijff 《The Journal of biological chemistry》1989,264(7):3988-3997
Monomolecular layers of lipid extracts of microsomal, mitochondrial outer and inner membranes, and pure lipid species have been used to measure their interaction with apo- and holocytochrome c. Large differences were observed both with respect to the nature and the lipid specificity of the interaction. The initial electrostatic interaction of the hemefree precursor apocytochrome c with anionic phospholipids is followed by penetration of the protein in between the acyl chains. Apocytochrome c shows similar interactions for all anionic lipids tested. In strong contrast the holoprotein discriminates enormously between cardiolipin for which it has a high affinity and phosphatidylserine and phosphatidylinositol for which it has a much lower affinity. For these latter lipids the interaction with cytochrome c is primarily electrostatic. The cytochrome c-cardiolipin interaction shows several unique features which suggest the formation of a specific complex between the two molecules. These properties account for the preference in interaction of the apoprotein with the lipid extract of the outer mitochondrial membrane over that of the endoplasmic reticulum and the large preference of cytochrome c for the inner over that of the outer mitochondrial membrane lipid extract. Only apocytochrome c was able to induce close contacts between monolayers of the mitochondrial outer membrane lipids and vesicles of mitochondrial inner membrane lipids. Experiments with fragments of both protein and unfolding experiments with cytochrome c revealed that the differences in interaction between the two proteins are mainly due to differences in their tertiary structure and not the presence of the heme group itself. The initial unfolded structure of apocytochrome c is responsible for the high penetrative power of the protein and its ability to induce close membrane contact, whereas the folded structure of cytochrome c is responsible for the specific interaction with cardiolipin. The results are discussed in the light of the apocytochrome c import process in mitochondria and suggest that lipid-protein interactions contribute to targeting the precursor toward mitochondria and are important for its translocation across the outer mitochondrial membrane and the final localization of cytochrome c toward the outside of the inner mitochondrial membrane. 相似文献