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1.
Aroclor 1254 was administered intraperitoneally (25 mg kg body wt −1 in 1 ml of arachis oil) at weekly intervals for 4 weeks to trout and carp; arachis oil was used as the control. Activities of the following hepatic microsomal enzymes, aminopyrine demethylase, p-nitroreductase, UDP-glucuronyl-transferase and 1-leucyl-β-naphthylamide splitting enzyme were measured in both species; cytochrome P 450 and microsomal protein contents were also determined. The changes in the levels of androgens, oestrogens and corticoid hormones were measured in the circulating blood of control and treated groups at weekly intervals. The blood was obtained by cardiac puncture. Results indicated (a) a significant increase in the activities of all the enzymes measured except 1-leucyl-β-naphthylamide splitting enzyme, (b) cytochrome P 450 and microsomal protein contents were increased in trout, but not in carp, (c) a significant reduction in the plasma levels of androgens, oestrogens and corticoids in the treated groups, particularly at the end of the fourth week and (d) there was a correlation between increased enzyme activities and a decrease in plasma hormone levels. 相似文献
2.
Abstract The presence of cytochrome P-450 and P-450-mediated benzo(a)pyrene hydroxylase activity in both microsomal and soluble fractions of the white rot fungus Phanerochaete chrysosporium was shown. The reduced carbon monoxide difference spectrum showed maxima at 448–450 and 452–454 nm for microsomal and cytosolic fractions, respectively. Both P-450 fractions produced a Type I substrate binding spectrum on addition of benzo(a)pyrene. Activity for benzo(a)pyrene hydroxylation was NADPH-dependent and inhibited by carbon monoxide. K m values for activity showed a difference between the cellular fractions with a K m of 89 μM for microsomal P-450 and 400 μM for cytosolic P-450. The V max values observed were 0.83 nmol min − (nmol microsomal P-450) −1 and 0.4 nmol min −1 (nmol cytosolic P-450) −1. The results indicate that P-450-mediated benzo(a)pyrene hydroxylase activity could play a role in xenobiotic transformation by this fungus beside the known ligninolytic exocellular enzymes. 相似文献
3.
Abstract A gene ( ERG11 ) encoding cytochrome P450 sterol 14α-demethylase (P450 14DM) was isolated from the maize pathogen, Ustilago maydis , by amplifying part of the coding region of the gene using PCR and by employing the amplified DNA fragment as a hybridization probe to recover the complete gene from an U. maydis λEMBL3 genomic library. The deduced amino acid sequence of the U. maydis gene showed homology to P450 14DMs from other organisms and contained specific motifs which were hallmarks of P450s. Expression of the gene in an U. maydis mutant (A20) deficient in P450 14DM led to only a partial restoration of P450 14DM activity. Accumulation of ergosta-7,22-dienol and ergost-7-enol in A20 transformants containing the ERG11 gene implied that an additional mutation affecting sterol Δ 5,6-desaturase activity accompanied the P450 14DM lesion. 相似文献
4.
Toxic Cu (II) effect on cytochrome b 559 under aerobic photoinhibitory conditions was examined in two different photosystem II (PSII) membrane preparations active in oxygen evolution. The preparations differ in the content of cytochrome b 559 redox potential forms. Difference absorption spectra showed that the presence of Cu (II) induced the oxidation of the high-potential form of cytochrome b 559 in the dark. Addition of hydroquinone reduced the total oxidized high-potential form of cytochrome b 559 present in Cu (II)-treated PSII membranes indicating that no conversion to the low-potential form took place. Spectroscopic determinations of cytochrome b 559 during photoinhibitory treatment showed slower kinetics of Cu (II) effect on cytochrome b 559 in comparison with the rapid loss of oxygen evolution activity in the same conditions. This result indicates that cytochrome b 559 is affected after PSII centres are photoinhibited. The high-potential form was more sensitive to toxic Cu (II) action than the low-potential form under illumination at pH 6.0. The content of the high-potential form of cytochrome b 559 was completely lost; however, the low-potential content was unaffected in these conditions. This loss did not involve cytochrome protein degradation. The results are discussed in terms of different binding properties of the heme iron to the protonated or unprotonated histidine ligand in the high-potential and low-potential forms of cytochrome b 559, respectively. 相似文献
5.
Abstract Cytochromes d and b 595 were studied by low temperature photodissociation of CO-ligated Azotobacter vinelandii membranes. White light or He-Ne laser irradiation revealed 436 and 594–597 nm absorption bands to be due to Fe 11 cytochrome b 595. Oxy-cytochrome d (648 nm) was formed when the CO adduct was photolysed in the presence of oxygen. This was followed by ligand recombination (presumably oxygen) to the high-spin cytochrome b 595, with a distinctive shift to shorter wavelengths of the α-band of the cytochrome, and a decrease in the oxygenated form. All spectral changes were light-reversible. We demonstrate the light-reversible binding of CO to both cytochromes b 595 and d , and suggest migration of oxygen from cytochrome d to cytochrome b 595 at a haem-haem binuclear centre during the oxidase reaction. 相似文献
6.
Bradyrhizobium japonicum utilizes cytochrome cbb 3 oxidase encoded by the fixNOQP operon to support microaerobic respiration under free-living and symbiotic conditions. It has been previously shown that, under denitrifying conditions, inactivation of the cycA gene encoding cytochrome c 550, the electron donor to the Cu-containing nitrite reductase, reduces cbb 3 expression. In order to establish the role of c 550 in electron transport to the cbb 3 oxidase, in this work, we have analyzed cbb 3 expression and activity in the cycA mutant grown under microaerobic or denitrifying conditions. Under denitrifying conditions, mutation of cycA had a negative effect on cytochrome c oxidase activity, heme c (FixP and FixO) and heme b cytochromes as well as expression of a fixP '–' lacZ fusion. Similarly, cbb 3 oxidase was expressed very weakly in a napC mutant lacking the c -type cytochrome, which transfers electrons to the NapAB structural subunit of the periplasmic nitrate reductase. These results suggest that a change in the electron flow through the denitrification pathway may affect the cellular redox state, leading to alterations in cbb 3 expression. In fact, levels of fixP '–' lacZ expression were largely dependent on the oxidized or reduced nature of the carbon source in the medium. Maximal expression observed in cells grown under denitrifying conditions with an oxidized carbon source required the regulatory protein RegR. 相似文献
7.
The compensatory growth of juvenile brown flounder Paralichthys olivaceus (body mass c. 12 g) following different thermal exposure was investigated. Fish were exposed to one of the five temperatures: 8·5 ( T 8·5), 13·0 ( T 13·0), 17·5 ( T 17·5), 22·0 ( T 22·0) and 26·5° C ( T 26·5) for 10 days and fish grew best at 22·0° C. Then the water temperature in all treatments was equably adjusted to 22·0° C over 3 days. At the end of the following 30 days after temperature adjustment, there were no significant differences between body masses of fish in the different treatments (wet body mass at the end of the experiment ranged from 22·13 to 24·56 g). Results indicated that the juvenile P. olivaceus achieved complete compensatory growth. Analysis of the dynamics of the feeding rates and feed conversion efficiencies indicated that compensatory growth of the fish experienced low temperature ( T 8·5, T 13·5 and T 17·5) or high temperature ( T 26·5) exposure was mainly dependent on increasing feed intake (hyperphagia) and possibly by improvement in feed conversion efficiency. The moisture content was not affected by different temperature exposure significantly. The lipid and energy content of juvenile P. olivaceus in T 8·5, however, were significantly lower than other treatment. Results of the current study indicate that a short period of low or high temperature exposure may not affect annual growth, but may affect lipid and energy deposition. 相似文献
8.
Abstract Cytochrome components which participate in the oxidation of nitrite in Nitrobacter winogradskyi have been highly purified and their properties studied in detail. Cytochrome a 1 c 1 is an iron-sulphur molybdoenzyme which has haems a and c and acts as a nitrite-cytochrome c oxidoreductase. Cytochrome c -550 is homologous to eukaryotic cytochrome c and acts as the electron mediator between cytochrome a 1 c 1 and aa 3-type cytochrome c oxidase. The oxidase is composed of two kinds of subunits, has two molecules of haem a and two atoms of copper in the molecule, and oxidizes actively eukaryotic ferrocytochrome c as well as its own ferrocytochrome c -550. Further, a flavoenzyme has been obtained which has transhydrogenase activity and catalyses reduction of NADP + with benzylviologen radical. This enzyme may be responsible for production of NADPH in N. winogradskyi . The electron transfer against redox potential from NO 2− to cythochrome c could be pushed through prompt removal by cytochrome aa 3 of H + formed by the dehydrogenation of NO 2−+ H 2O. As cytochrome c in anaerobically kept cell-free extracts is rapidly reduced on addition of NO 2−, a membrane potential does not seem necessary for the reduction of cytochrome c by cytochrome a 1 c 1 with NO 2− in vivo. 相似文献
9.
Abstract The role of cytochrome b 562, a fragile constituent of the respiratory terminal oxidase supercomplex of the thermoacidophilic archaeon, Sulfolobus sp. strain 7, was investigated spectroscopically in the membrane-bound state. Cytochrome b 562 did not react with CO or cyanide in the membrane-bound state, while it was irreversibly modified to a CO-reactive form ( b 562) upon solubilization in the presence of cholate and LiCl. Cyanide titration analyses with the succinate-reduced membrane suggested that cytochrome b 562 was upstream of both the ' g y= 1.89' Rieske FeS cluster and the a -type cytochromes. These results show that the b -type cytochrome functions as an intermediate electron transmitter in the terminal oxidase supercomplex. 相似文献
10.
Experiments were carried out to evaluate the effects of exposure to nitric oxide on the ability by NADPH‐dependent microsomal electron transfer to generate oxygen radicals. Such interactions could play a role in the potential antioxidant action of nitric oxide (NO). Isolated microsomes from soybean ( Glycine max [L.] Merr. cv. Hood) embryonic axes were exposed to an exogenously added source of nitric oxide (NO) (S‐nitrosoglutathione + dithiothreitol). The O 2− generation rate by microsomes exposed to NO decreased significantly as compared to the rate measured in microsomes incubated in the absence of NO. The exposure of the microsomes to the NO donor did not alter the microsomal rate of hydroxyl radical generation. Preincubation of the microsomes with the NO donor affected neither iron reduction rate nor activity of cytochrome c reductase. However, cytochrome P 450 activity was significantly inhibited after exposure to NO. This inhibition was completely prevented by hemoglobin. The data are consistent with the hypothesis that NO exhibits a potential antioxidant role in the plant cell by decreasing the rate of generation of superoxide anion. Since endogenous NO was detected in homogenates of soybean embryonic axes by EPR studies, this interaction between NO and cytochrome P 450 in soybean embryonic axes could be a factor of relevance for the control of oxidative stress in vivo. 相似文献
11.
The content of cytochrome P-450 and the specific activity of laurate hydroxylation have been measured at monthly intervals through the year in liver microsomes of males and females of Japanese bullfrog, Rana catesbeiana. The specific activity, based on microsomal protein or cytochrome P-450 of both sexes was higher in spring (April-May) than in autumn (October-November) and the difference was statistically significant. However, in males the content of the cytochrome P-450 in the spring was almost the same level as in the autumn and it was rather lower in the spring than in the autumn in females. These suggest that the cytochrome P-450 species catalyzing laurate hydroxylation increased specifically in the spring. The specific activity (based on cytochrome P-450) in males was significantly higher than that in females in spring (April-May) or autumn (October-November). 相似文献
12.
The population structure and seasonal changes in condition factors of the burbot in a shallow coastal region of the north-eastern Bothnian Bay are described. The significance of the so-called rest years is examined by comparing condition indices in immature or sterile and mature burbot. The somatic condition index ( K 2), liver index ( K 1), intestine index ( K 1) and gonad index ( K g ) are determined monthly in terms of organ weight in relation to body length. Approximately 30% of the whole catch of 1052 burbot were non-maturing but were 40 cm or more in length. K 2, K 1 and K 1 were lowest in the autumn, when the first sign of gonad recrudescence was observed in mature burbot. The non-maturing burbot were never in poorer condition than mature ones. As mature and non-maturing burbot dissipated their energy stores during the warmest period of the summer, it is concluded that burbot spending a rest year do not accumulate and store energy reserves over the summer for the next year, and that such rest years, if they exist, do not occur for nutritional reasons. 相似文献
13.
DNA from archived otoliths was used to explore the temporal stability of the genetic composition of two cod populations, the Moray Firth (North Sea) sampled in 1965 and 2002, and the Bornholm Basin (Baltic Sea) sampled in 1928 and 1997. We found no significant changes in the allele frequencies for the Moray Firth population, while subtle but significant genetic changes over time were detected for the Bornholm Basin population. Estimates of the effective population size ( N e ) generally exceeded 500 for both populations when employing a number of varieties of the temporal genetic method. However, confidence intervals were very wide and N e 's most likely range in the thousands. There was no apparent loss of genetic variability and no evidence of a genetic bottleneck for either of the populations. Calculations of the expected levels of genetic variability under different scenarios of N e showed that the number of alleles commonly reported at microsatellite loci in Atlantic cod is best explained by N e 's exceeding thousand. Recent fishery-induced bottlenecks can, however, not be ruled out as an explanation for the apparent discrepancy between high levels of variability and recently reported estimates of N e << 1000. From life history traits and estimates of survival rates in the wild, we evaluate the compatibility of the species' biology and extremely low N e / N ratios. Our data suggest that very small N e 's are not likely to be of general concern for cod populations and, accordingly, most populations do not face any severe threat of losing evolutionary potential due to genetic drift. 相似文献
15.
Bradyrhizobium japonicum cytochrome c 550, encoded by cycA , has been previously suggested to play a role in denitrification, the respiratory reduction of nitrate to dinitrogen. However, the exact role of this cytochrome in the denitrification process is unknown. This study shows that cytochrome c 550 is involved in electron transfer to the copper-containing nitrite reductase of B. japonicum , as revealed by the inability of a cycA mutant strain to consume nitrite and, consequently, to grow under denitrifying conditions with nitrite as the electron acceptor. Mutation of cycA had no apparent effect on methylviologen-dependent nitrite reductase activity. However, succinate-dependent nitrite reduction was largely inhibited, suggesting that c 550 is the in vivo electron donor to copper-containing nitrite reductase. In addition, this study demonstrates that a cytochrome c 550 mutation has a negative effect on expression of the periplasmic nitrate reductase. This phenotype can be rescued by extending the growth period of the cells. A model is proposed whereby a mutation in cycA reduces expression of the cbb 3-type oxidase, affecting oxygen consumption rate by the cells and consequently preventing maximal expression of the periplasmic nitrate reductase during the first days of the growth period. 相似文献
16.
The physiological and anaesthetic effects of three different levels of air-saturated and buffered CO 2 anaesthesia, P w co 2=31, 78, or 125 mmHg, were examined in cannulated rainbow trout Oncorhynchus mykiss . Complete anaesthesia (no opercular movements) was not achieved by these hypercapnic levels after 20 min of CO 2 exposure. Although increasing P w co 2 reduced the induction times to the early stages of anaesthesia, it also resulted in increasing hyperventilatory, hypoxaemic, and acid-base disturbances. After a 10-min recovery period, while the respiratory acidosis component of the acid-base disturbance was corrected, there was a significant metabolic acidosis. Recovery time was longest in the high P w co 2 treatment where 33% of the fish died. Two additional groups ( P w co 2=37 and 78 mmHg) were exposed to an acute stress prior to the anaesthetic treatment. Stress reduced the hypoventilatory effects of the low P w co 2 treatment, increased the recruitment of anaerobic metabolism, and prolonged recovery time. Although the increase in plasma catecholamines elicited by the stress was small relative to the response obtained with the anaesthetic, stress prior to CO 2 anaesthesia impaired the efficiency of the treatment. Overall, our results suggest that P w co 2 levels above 37 mmHg and/or stress prior to the anaesthesia impair the efficiency of air-saturated and buffered CO 2 anaesthesia by exacerbating the hypoxaemic effects of the hypercapnic treatment. 相似文献
17.
The biochemical basis for the complex effects of the anti-cancer drug cisplatin on hepatic cytochrome P-450 activity was studied in adult male rat liver using P-450 form-specific steroid hydroxylase assays and antibody probes. Cisplatin treatment of adult male rats resulted in a marked and prolonged feminization of the pattern of P-450 enzymes expressed in hepatic tissue. The adult male-specific cytochrome P-450 forms designated P-450 2c (P-450 gene IIC11), P-450 2a (gene IIIA2), and P-450 RLM2 were decreased by 70-90% after 7-14 days, with parallel decreases in their respectively associated microsomal steroid hydroxylase activities. Concomitantly, hepatic levels of the female-predominant enzymes P-450 3 (gene IIA1) and P-450j (gene IIE1) were elevated approximately 2-4-fold. The female-specific microsomal enzyme androstenedione 5 alpha-reductase was induced approximately 20-fold by cisplatin; however, no elevation of the female-specific P-450 2d was detected. The underlying hormonal basis for these effects of cisplatin was then examined. Serum testosterone levels were found to be depleted by cisplatin in a time- and dose-dependent manner which correlated with the observed changes in these hepatic enzymes. Furthermore, castration of adult rats altered the profile of these enzymes in a manner which resembled that observed with cisplatin treatment, suggesting that androgen depletion was the primary cause for the observed feminization of hepatic enzyme expression. Consistent with this possibility, the synthetic androgen methyltrienolone effectively blocked the changes in hepatic enzyme expression induced by cisplatin. Moreover, hepatic enzyme feminization was significantly reversed by chorionic gonadotropin, which fully restored serum testosterone levels in the cisplatin-treated rat. Luteinizing hormone-releasing hormone challenge experiments demonstrated that the responsiveness of the pituitary to this hypothalamic regulator of testicular androgen production was unimpaired by cisplatin treatment, indicating that hypothalamic production or secretion of luteinizing hormone-releasing hormone may be deficient in the cisplatin-treated animals. These studies establish that the effects of cisplatin on hepatic P-450 enzyme expression result from its interruption of the hypothalamic-pituitary stimulation of testicular androgen production and that this, in turn, leads to a depletion of circulating androgens required for maintenance of normal P-450 enzyme expression in adult male rats. 相似文献
18.
Abstract Evidence is presented to show that the thiosulphate-oxidising multi-enzyme system from Thiobacillus versutus has a periplasmic location, and that the oxygen-binding site of the cytochrome oxidase ( aa 3) is on the inner surface of the membrane. A scheme for the mechanism of generation of a proton motive force during electron flow from thiosulphate to oxygen via cytochrome c and aa 3 is proposed. 相似文献
19.
Androgen aromatase was found to also be estrogen 2-hydroxylase. The substrate specificity among androgens and estrogens and multiplicity of aromatase reactions were further studied. Through purification of human placental microsomal cytochrome P-450 by monoclonal antibody-based immunoaffinity chromatography and gradient elution on hydroxyapatite, aromatase and estradiol 2-hydroxylase activities were co-purified into a single band cytochrome P-450 with approx. 600-fold increase of both specific activities, while other cytochrome P-450 enzyme activities found in the microsomes were completely eliminated. The purified P-450 showed M r of 55 kDa, specific heme content of 12.9 ± 2.6 nmol·mg −1 (±SD, N = 4), reconstituted aromatase activity of 111 ± 19 nmol·min −1·mmg −1 and estradiol 2-hydroxylase activity of 5.85 ± 1.23 nmol·min −1·mg −1. We found no evidence for the existence of catechol estrogen synthetase without concomitant aromatase activity. The identity of the P-450 for the two different hormone synthetases was further confirmed by analysis of the two activities in the stable expression system in Chinese hamster ovarian cells transfected with human placental aromatase cDNA, pH β-Aro. Kinetic analysis of estradiol 2-hydroxylation by the purified and reconstituted aromatase P-450 in 0.1 M phosphate buffer (pH 7.6) showed Km of 1.58 μM and Vmax of 8.9 nmol·min −1·mg −1. A significant shift of the optimum pH and Vmax, but not the Km, for placental estrogen 2-hydroxylase was observed between microsomal and purified preparations. Testosterone and androstenedione competitively inhibited estradiol 2-hydroxylation, and estrone and estradiol competitively inhibited aromatization of both testosterone and androstenedione. Estrone and estradiol showed Ki of 4.8 and 7.3 μM, respectively, for testosterone aromatization, and 5.0 and 8.1 μM, respectively, for androstenedione aromatization. Androstenedione and testosterone showed Ki of 0.32 and 0.61 μM, respectively, for estradiol 2-hydroxylation. Our studies showed that aromatase P-450 functions as estrogen 2-hydroxylase as well as androgen 19-, 1β-,and 2β-hydroxylase and aromatase. The results indicate that placental aromatase is responsible for the highly elevated levels of the catechol estrogen and 19-hydroxyandrogen during pregnancy. These results also indicate that the active site structure holds the steroid ssubstrates to face their β-side of the A-ring to the heme, tilted in such a way as to make the 2-position of estrogens and 19-, 1-, and 2-positions of androgens available for monooxygenation. 相似文献
20.
Aromatase is a microsomal cytochrome P450 that converts androgens to estrogens by three sequential oxidations. The isolation of the 19-hydroxy and 19-oxo androgens suggests that the first two oxidations occur at the C 19 carbon. However, the mechanism of the third oxidation, which results in C 10---C 19 bond cleavage, has not been determined. Two proposed mechanisms which remain viable involve either initial 1β-hydrogen atom abstraction or addition of the ferric peroxy anion from aromatase to the C 19 aldehyde. Semiempirical molecular orbital calculations (AM1) were used to study potential reaction mechanisms initiated by initial 1β-hydrogen atom abstraction. Initially, the energetics of carbon---carbon bond cleavage of the keto and enol forms of C1-radicals were studied and were found to be energetically similar. A mechanism was proposed in which the 19-oxo intermediate is subject to initial nucleophilic attack by the protein. The geometry of the A-ring in the androgens is between that for the 1-radicals and estrogen, suggesting that some transition state stabilization for the homolytic cleavage reaction can occur. More recently, studies on liver microsomal cytochrome P450 mediated deformylation of xenobiotic aldehydes supports mechanisms involving an alkyl peroxy intermediate formed by addition of the ferric peroxy anion from aromatase to the C19 aldehyde. Although this intermediate could proceed through several different concerted or non-concerted pathways, one non-concerted pathway involves the heterolytic cleavage of the dioxygen bond resulting in an active oxygenating species (iron-oxene) and a diol. The diol could then undergo hydrogen atom abstraction followed by homolytic carbon---carbon bond cleavage as in the mechanisms modeled previously. When this cleavage was modeled for seven aldehydes, a good correlation with reported experimental aldehyde turnover numbers was obtained. However, when dialkoxy derivatives of the aldehydes are subject to microsomal metabolism, the rates of carbon---carbon cleavage products do not approach the rates of deformylation of the aldehyde analog. 相似文献
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