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1.
The midgut of Hyalophora cecropia actively transports potassium from hemolymph to lumen and the energy for this process appears to be intimately linked to oxidative metabolism. In the present investigation, we monitored concurrently the rate of active transport and the redox levels of the components of the respiratory chain in the intact tissue under a variety of experimental conditions. Approximately equal concentrations of cytochromes a3, a, c and b-557 were found. Other investigators (Pappenheimer, Jr, A.M. and Williams, C.M. (1954) J. Biol. Chem. 209, 915, Shappirio, D.G. and Williams, C.M. (1957) Proc. R. Soc. Lond. Ser. B 147, 233 and Chance, B. and Pappenheimer, Jr, A.M. (1957) J. Biol, Chem, 209, 931) have indentified cytochrome b-557 with b5 and found that it exists primarily in an extramitochondrial location. Steady-state experiments demonstrated that all these cytochromes were approximately 50% reduced while active transport proceeded at a high rate in regular cecropia Ringer containing 32 mM KCl. When the potassium concentration was reduced, the active transport decreased and all the cytochromes became more oxidized. Addition of 1 mM cyanide inhibited active transport by 90% and caused a 100% reduction of all cytochromes. Redox state and short circuit current (Isc) kinetics measured as the tissue was made anoxic showed that all the respiratory enzymes, except cytochrome b-557, became fully reduced at a faster rate than the rate of inhibition of the Isc. The rate of cytochrome b-557 reduction followed kinetically the Isc. These observations are interpreted in a scheme where cytochrome b-557 (possibly b5) branches off cytochrome c from the conventional resporatory chain, utilizing cytochrome a3 as the terminal oxidase for both branches. Cytochrome b-557 may be involved in providing a direct link between oxidative metabolism and active transport in the midgut of the silkworm. 相似文献
2.
The respiratory pigments of Prototheca zopfii include seven cytochromes: two c-type cytochromes, a soluble c(549) and a membrane bound c(551); three b-type cytochromes, b(555), b(559) and b(564); and cytochromes a and a3. Cytochromes a and a3 could be resolved spectrally in the α-band region by reducing the cells in the presence of methanol and cyanide. Methanol shifted the absorption maximum of cytochrome a from 598 to 603 nanometers and permitted dithionite (or substrate) to reduce the cyanide-cytochrome a3 complex to give a well defined 595-nanometer absorption band. Methanol did not interfere with CO binding by cytochrome a3, and CO did not alter the methanol effect on cytochrome a. Azide and cyanide, which partially inhibited exogenous respiration, stimulated endogenous respiration. Frozen steady states of the electron transport chain in the presence of cyanide and azide indicated that the stimulation by these inhibitors was due to an increased autooxidation of one of the b-type cytochromes, possibly b(564). 相似文献
3.
A minor form of hepatic microsomal cytochrome P-450 has been purified to apparent homogeneity from rats treated with the polychlorinated biphenyl mixture, Aroclor 1254. This newly isolated hemoprotein, cytochrome P-450 e, is inducible in rat liver by Aroclor 1254 and phenobarbital, but not by 3-methylcholanthrene. Two other hemoproteins, cytochromes P-450 b and P-450 c, have also been highly purified during the isolation of cytochrome P-450 e based on chromatographic differences among these proteins. By Ouchterlony double-diffusion analysis with antibody to cytochrome P-450 b, highly purified cytochrome P-450 e is immunochemically identical to cytochrome P-450 b but does not cross-react with antibodies prepared against other rat liver cytochromes P-450 ( P-450 a, P-450 c, P-450 d) or epoxide hydrolase. Purified cytochrome P-450 e is a single protein-staining band in sodium dodecyl sulfate-polyacrylamide gels with a minimum molecular weight (52,500) slightly greater than cytochromes P-450 b or P-450 d (52,000) but clearly distinct from cytochromes P-450 a (48,000) and P-450 c (56,000). The carbon monoxide-reduced difference spectral peak of cytochrome P-450 e is at 450.6 nm, whereas the peak of cytochrome P-450 b is at 450 nm. Ethyl isocyanide binds to ferrous cytochromes P-450 e and P-450 b to yield two spectral maxima at 455 and 430 nm. At pH 7.4, the 455:430 ratio is 0.7 and 1.4 for cytochromes P-450 b and P-450 e, respectively. Metyrapone binds to reduced cytochromes P-450 e and P-450 b (absorption maximum at 445–446 nm) but not cytochromes P-450 a, P-450 c, or P-450 d. Metabolism of several substrates catalyzed by cytochrome P-450 e or P-450 b reconstituted with NADPH-cytochrome c reductase and dilauroylphosphatidylcholine was compared. The substrate specificity of cytochrome P-450 e usually paralleled that of cytochrome P-450 b except that the rate of metabolism of benzphetamine, benzo[ a]pyrene, 7-ethoxycoumarin, hexobarbital, and testosterone at the 16α-position catalyzed by cytochrome P-450 e was only 15–25% that of cytochrome P-450 b. In contrast, cytochrome P-450 e catalyzed the 2-hydroxylation of estradiol-17β more efficiently (threefold) than cytochrome P-450 b. Cytochrome P-450 d, however, catalyzed the metabolism of estradiol-17β at the greatest rate compared to cytochromes P-450 a, P-450 b, P-450 c, or P-450 e. The peptide fragments of cytochromes P-450 e and P-450 b, generated by either proteolytic or chemical digestion of the hemoproteins, were very similar but not identical, indicating that these two proteins show minor structural differences. 相似文献
4.
The electron transport systems of Fasciola hepatica mitochondria were investigated spectrophotometrically at room temperature and at −196°. The mitochondria were found to contain substrate reducible a-, b- and c-type cytochromes. All of the cytochrome components of the classical mammalian type of respiratory chain were present, although the concentration of cytochromes aa3 was low. In addition to the mammalian type of respiratory chain, the Fasciola mitochondria contained a substrate reducible b-type cytochrome component (557 nm) which included a CO reactive o-type cytochrome. The results suggest that F. hepatica mitochondria contain a branched electron transport system including a mammalian type of chain and involving two terminal oxidases and at least two b-type cytochromes. 相似文献
6.
The effect of Ba 2+ on Na + transport and electrical characteristics of toad bladder was determined from change produced in short circuit current ( Isc, epithelial, apical and basal-lateral potentials ( ψt, ψa, ψb), epithelial and membrane resistances ( Rt, Ra, Rb) and shunt resistance ( Rs). Mucosal Ba 2+ had no effect. Serosal Ba 2+ reduced Isc, ψt, ψa, and ψb, but had no effect on Rt, Ra, Rb and Rs. Minimal effective Ba 2+ concentration was 5 · 10 ?5 M. The phenomenon was reversed by Ba 2+ removal, but not by 86 mM serosal K +. Ba 2+ inhibition of Isc did not impair the response to vasopressin which was quantitatively the same as controls. ψa with Ba 2+ equalled ψb. After Ba 2+ inhibition, ouabain produced no further decrease in ψt and Isc. Ba 2+ exposure after ouabain did not decrease ψt and Isc. The results suggest that Ba 2+ inhibits the basal-lateral electrogenic Na + pump. 相似文献
7.
The cytochrome complement of Methylophilus methylotrophus and its respiratory properties were determined during batch culture and in continuous culture under conditions of methanol-, nitrogen- and O 2-limitation. About 35% of the cytochrome c produced by the bacteria was released into the growth medium, and of the remaining cytochrome c about half was membrane-bound and half was soluble. Two cytochromes c were always present on membranes (redox potentials 375mV and 310mV), and these probably correspond to the soluble cytochromes c described previously [Cross & Anthony (1980) Biochem. J. 192, 421–427]. A third minor component of cytochrome c (midpoint potential 356mV) was only detected on membranes of methanol-limited bacteria. M. methylotrophus always contained two membrane-bound cytochromes b with α-band absorption maxima of about 556 and 563nm (measured at room temperature) and midpoint potentials of 110 and 60mV respectively. There appeared to be relatively more of the cytochrome b563 in methanol-limited bacteria. A third b-type cytochrome with an α-band absorption maximum at 558 (at 77K) reacted with CO and had a high midpoint redox potential (260mV); it is thus a potential oxidase and hence is called cytochrome o. The roles of these cytochromes in electron transport were confirmed by investigating the patterns of respiratory inhibition. It is proposed that two cytochromes are physiological oxidases: cytochrome a+ a3, which is present only in methanol-limited conditions, and the cytochrome o, which is induced 10-fold in conditions of methanol excess. Schemes for electron transport from methanol and NAD(P)H to O 2 in M. methylotrophus under various limitations are proposed. Spectra and potentiometric titrations of cytochromes in whole cells and membranes of M. methylotrophus grown under various nutrient limitations have been deposited as Supplementary Publication SUP 50111 (10 pages) at the British Library Lending Division, Boston Spa, Wetherby, West Yorkshire LS23 7BQ, U.K., from whom copies can be obtained on the terms indicated in Biochem. J. (1978) 169, 5. 相似文献
10.
In highly aerobic environments, Gluconacetobacter diazotrophicus uses a respiratory protection mechanism to preserve nitrogenase activity from deleterious oxygen. Here, the respiratory system was examined in order to ascertain the nature of the respiratory components, mainly of the cyanide sensitive and resistant pathways. The membranes of G. diazotrophicus contain Q 10, Q 9 and PQQ in a 13:1:6.6 molar ratios. UV 360 nm photoinactivation indicated that ubiquinone is the electron acceptor for the dehydrogenases of the outer and inner faces of the membrane. Strong inhibition by rotenone and capsaicin and resistance to flavone indicated that NADH-quinone oxidoreductase is a NDH-1 type enzyme. KCN-titration revealed the presence of at least two terminal oxidases that were highly sensitive and resistant to the inhibitor. Tetrachorohydroquinol was preferentially oxidized by the KCN-sensitive oxidase. Neither the quinoprotein alcohol dehydrogenase nor its associated cytochromes c were instrumental components of the cyanide resistant pathway. CO-difference spectrum and photodissociation of heme-CO compounds suggested the presence of cytochromes b-CO and a1-CO adducts . Air-oxidation of cytochrome b (432 nm) was arrested by concentrations of KCN lower than 25 μM while cytochrome a1 (442 nm) was not affected. A KCN-sensitive ( I50 = 5 μM) cytochrome bb and a KCN-resistant ( I50 = 450 μM) cytochrome ba quinol oxidases were separated by ion exchange chromatography. 相似文献
11.
1. Purified mitochondria have been prepared from wild type Paramecium tetraurelia and from the mutant Cl 1 which lacks cytochrome aa3. Both mitochondrial preparations are characterized by cyanide insensitivity. Their spectral properties and their redox potentials have been studied.2. Difference spectra (dithionite reduced minus oxidized) of mitochondria from wild type P. tetraurelia at 77 K revealed the α peaks of b-type cytochrome(s) at 553 and 557 nm, of c-type cytochrome at 549 nm and a-type cytochrome at 608 nm. Two α peaks at 549 and 545 nm could be distinguished in the isolated cytochrome c at 77 K. After cytochrome c extraction from wild type mitochondria, a new peak at 551 nm was unmasked, probably belonging to cytochrome c1. The a-type cytochrome was characterized by a split Soret band with maxima at 441 and 450 nm. The mitochondria of the mutant Cl 1 in exponential phase of growth differed from the wild type mitochondria in that cytochrome aa3 was absent while twice the quantity of cytochrome b was present. In stationary phase, mitochondria of the mutant were characterized by a new absorption peak at 590 nm.3. Cytochrome aa3 was present at a concentration of 0.3 nmol/mg protein in wild type mitochondria and ubiquinone at a concentration of 8 nmol/mg protein both in mitochondria of the wild type and the mutant Cl 1. Cytochrome aa3 was more susceptible to heat than cytochromes b and c, c1.4. CO difference spectra at 77 K revealed two different Co-cytochrome complexes. The first, found only in wild type mitochondria, was a typical CO-cytochrome a3 complex characterized by peaks at 596 and 435 nm and troughs at 613 and 450 nm. The second, found both in mitochondria of the wild type and the mutant, was a CO-cytochrome b complex with peaks at 567, 539 and 420 nm and a trough at 558-549 nm. Both complexes are photo-dissociable.5. Spectral evidence was obtained for interaction of cyanide with the a-type cytochrome (shift of the α peak at 77 K from 608 to 605 nm), but not with the b-type cytochrome.6. The mid-point potentials of the different cytochromes at neutral pH are as follows: cytochrome aa3 235 and 395 mV, cytochrome c, c1 233 mV, cytochromes b 120 mV. 相似文献
12.
(1) Cells of ThiobacillusA2 grown chemoautotrophically on thiosulfate or heterotrophically on succinate with oxygen contained b-, c-, o-, a- and a3-type cytochromes. The amount of cytochrome per mg of cell protein was much greater in thiosulfate-grown cells and differences in the relative concentrations of cytochromes were observed for the different growth conditions. (2) The half-reduction potentials at pH 7.0 ( Em,7.0) and spectral maxima of c-, b-, a- and a3-type cytochromes were similar in cells grown aerobically with thiosulfate or with succinate as the growth substrate. (3) The half-reduction potential of the ‘invisible’, or high-potential copper, as determined from the potentiometric behavior of the carbon monoxide-reduced cytochrome a3 complex at pH 8.0, was 365 mV. (4) Reducing equivalents from thiosulfate appear to enter the respiratory chain at the cytochrome c level; however, studies in cell-free extracts were limited due to a loss in respiratory activity with thiosulfate as a substrate upon cell disruption. 相似文献
13.
The rate of incorporation of [ 14C]aminolevulinic acid (ALA) into cytochrome hemes was used to measure mitochondrial cytochrome synthesis in the fat body of adult male Blaberus discoidalis cockroaches. The hemes of cytochromes aa3+ b and c+ c1, were chemically separated to observe differential rates in their synthesis and regulation. [ 14C]ALA was linearly incorporated into cytochrome hemes for at least 8 h. No significant pool of endogenous ALA was detected relative to the amount of administered [ 14C]ALA. Peak cytochrome synthesis occurred 4 to 6 days after adult emergence. Endocrine disruption by corpora cardiaca-corpora allata extirpation or cervical ligation eliminated the 4-day developmentally related increase in the rate of cytochrome aa3+ b synthesis but had no effect on the production of cytochromes c+ c1. Injections of corpora cardiaca extracts into cervically ligated animals stimulated the rate of production of cytochromes aa3+ b by 2.5 times but did not affect cytochromes c+ c1. By comparison, juvenile hormone injections did not affect the rate of synthesis of either cytochrome fraction. These findings indicate that a neurohormone regulates the rate of synthesis of cytochromes a+ b in insect fat body mitochondria. 相似文献
14.
1. Mitochondria of the obligately aerobic ciliate protozoon, Tetrahymena pyriformis strain ST, are unusual in that they possess a cytochrome oxidase system that does not react with reduced mammalian cytochrome c; the presence of cytochromes a603+ a3 is masked in the α-band region of spectra by the broad absorption band of cytochrome a620. 2. Other haemoproteins present include cytochromes b560, b556, c553 and c549. 3. The reaction of reduced cytochrome a3 with CO is reversed by flash photolysis, and in the presence of O 2 the subsequent oxidation of this cytochrome is followed by that of cytochrome a603. 4. Cytochromes a620 and b560 also react with CO and with KCN; the latter cytochrome corresponds with that designated cytochrome o by other workers. 5. The contribution of cytochrome a603 to difference spectra is revealed by making use of the fact that it does not react with KCN. 6. Cytochrome a620 is unstable, and its α-absorption band is lost from spectra of mitochondria which have been aged or treated with ultrasound, detergents or organic solvents. 7. Possible pathways of electron transport via the several different terminal oxidases in Tetrahymena mitochondria are proposed. 相似文献
15.
Tightly coupled mitochondria have been prepared from a variety of plant sources: white potato ( Solanum tuberosum), Jerusalem artichoke ( Heliantus tuberosus), cauliflower buds ( Brassica oleracea), and mung bean hypocotyls ( Phaseolus aureus). Mitochondria with no appreciable coupling were also prepared from skunk cabbage spadices ( Symplocarpus foetidus). Room temperature difference spectra show that these mitochondria are very similar in the qualitative and quantitative composition of their electron carriers. The different cytochromes are present in the amounts of 0.1 to 0.3 mμmole per mg of mitochondrial protein. The molar ratios of the different electron carriers are, on the average: 0.7:0.7:1.0:3 to 4:10 to 15 respectively for cytochrome aa3, cytochromes b, cytochromes c, flavoproteins, and pyridine nucleotides. From low temperature difference spectra carried out under particular experimental conditions, it can be deduced that these mitochondria contain 3 b cytochromes whose α bands are located at 552, 557, and 561 mμ, and 2 c cytochromes, one of which, a c1-like cytochrome, is firmly bound to the mitochondrial membrane. Cytochrome oxidase can be optically resolved into its 2 components a and a3. For all kinds of mitochondria, the rates of oxidation of succinate are similar as well as the turnover of cytochrome oxidase (50-70 sec−1), regardless of the metabolic activities of the tissues. The number of mitochondria per cell appears to be the controlling factor of the intensity of tissue respiration. 相似文献
16.
Irradiation of cells of Prototheca zopfii with blue light inhibited the respiratory capacity of the cells. The inhibition of respiration was correlated with a photodestruction of cytochrome c(551), cytochrome b(559), and cytochrome a3. Cytochrome c(549), cytochrome b(555), and cytochrome b(564) were unaffected by the irradiation treatment. The α-band of reduced cytochrome a was shifted from 599 to 603 nm by irradiation, an effect similar to that observed when methanol was added to nonirradiated cells. The presence of oxygen was required during irradiation for both photoinhibition of respiration and photodestruction of the cytochromes. Cytochrome a3 was protected against photodestruction by cyanide. Photodestruction of these same cytochromes also occurred when washed mitochondria of P. zopfii were irradiated. 相似文献
18.
1. The participation of cytochromes in the membrane-bound, nitrate and oxygen respiratory systems of Klebsiella (Aerobacter) aerogenes has been investigated. The membrane preparations contained the NADH, succinate, lactate and formate oxidase systems, and in addition a high respiratory nitrate reductase activity.2. Difference spectra indicated the presence of cytochromes b, a1, d, and o. Cytochromes of the c-type could not be detected in these membranes. Both cytochrome b content and respiratory nitrate reductase activity were the highest in bacteria grown anaerobically in the presence of nitrate.3. Cytochrome b was the only cytochrome which, after being reduced by NADH, could be partially reoxidized anaerobically in the presence of nitrate. Furthermore, nitrate caused a lower aerobic steady state reduction only of cytochrome b.4. NADH oxidase and NADH-linked respiratory nitrate reductase activities were both inhibited by antimycin A, 2- n-heptyl-4-hydroxyquinoline- N-oxide and KCN. NADH oxidase activity was selectively inhibited by CO, while azide was found to inhibit only the respiratory nitrate reductase. In the presence of azide, nitrate did not affect the level of reduction of cytochrome b.5. The evidence presented suggests that cytochrome b is a carrier in the electron transport systems to both nitrate and oxygen; from cytochrome b branching occurs, with one branch linked to the respiratory nitrate reductase and one branch linked to oxidase systems, containing the cytochromes a1, d and o. 相似文献
19.
Cytochromes b553, b557, and b562 of mung bean ( Phaseolus aureus) mitochondria become partially reduced with endogenous substrate on addition of antimycin A to the aerobic mitochondrial suspension. Addition of ATP causes partial reoxidation of the three cytochromes. This partial oxidation by ATP is inhibited by oligomycin and reversed by uncoupler. Ubiquinone does not appear to act as electron acceptor for the oxidation reaction, but a nonfluorescent flavoprotein, or possibly ironsulfur protein, component does appear to act as acceptor. This is consistent with reverse electron transport driven by ATP across the first site of energy conservation of the respiratory chain. Endogenous pyridine nucleotide and the fluorescent flavoprotein with E m7.2 = −155mv (midpoint potential at pH 7.2, referred to normal hydrogen electrode) in uncoupled mitochondria become reduced in anaerobiosis attained by oxidation of succinate in the absence of respiratory inhibitors of the cytochrome chain, provided that Pi and ATP are present. Under these same conditions, cytochrome b557 is completely reduced but cytochrome b562 remains nearly completely oxidized. There is no equilibration across the first site of energy conservation between the carriers on the low potential side and cytochrome b562 with E m7.2 = −77mv on the high potential side. It is concluded that cytochrome b562 is not a part of the main sequence of electron transport carriers of the mitochondrial respiratory chain of plants; it can participate in redox reactions with the respiratory chain in coupled mitochondria but not in uncoupled mitochondria unless antimycin A is present. 相似文献
20.
The electron transport systems of 2 species of aquatic fungi, Leptomitus lacteus and A podachlya punctata, contained cytochrome a-a3 (605 mμ), 2 b type cytochromes (564 and 557 mμ), c type cytochrome (551 mμ), and flavoprotein, but they appeared to lack cytochrome c1. Reduced-minus-oxidized difference spectra and difference spectra in the presence of antimycin A or cyanide were used to characterize these systems. Studies with the electron microscope revealed that hyphae of Leptomitus lacteus contained numerous, conspicuous mitochondria with tubular cristae. 相似文献
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