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41.
Fate of corticotrophins in an isolated adrenal-cell bioassay and decrease of peptide breakdown by cell purification 总被引:5,自引:3,他引:2
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Hugh P. J. Bennett Gillian Bullock P. J. Lowry Colin McMartin Judith Peters 《The Biochemical journal》1974,138(2):185-194
1. The fate of corticotrophins in a trypsin-dispersed rat adrenal-cell assay system was investigated with a view to establishing whether differences in the rate of inactivation might contribute to potency differences observed between analogues. 2. Corticotrophin-(1-24)-tetracosapeptide and to a lesser extent synthetic 1-39 corticotrophins were found to be inactivated during incubation with cell suspension. 3. Peptide fragments were isolated by using [[(3)H(2)]Tyr(23)]corticotrophin-(1-24)- tetracosapeptide as a marker. The fragments indicate a peptidase with a predominantly tryptic specificity. 4. The peptidase is present in the extracellular fluid and is released from cells when they are damaged. 5. Cells were fractionated on an albumin gradient. Cells from the zona fasciculata and the zona intermedia or reticularis were present in fractions which produced fluorogenic steroids in response to corticotrophin. 6. Purification of the cells by centrifugation through albumin decreased degradation by peptidases, so that if the assay is carried out with a dilute suspension of purified cells peptide breakdown should not affect the observed potencies of adrenocorticotrophin analogues. 7. No binding of [[(3)H(2)]Tyr(23)]corticotrophin-(1-24)- tetracosapeptide to cells could be detected at low concentrations of the peptide. This indicated that less than 120 receptors/cell are occupied during stimulation by a dose that would elicit approx. 80% of the maximal response. 相似文献
42.
Studies on partially reduced mammalian cytochrome oxidase. Reactions with carbon monoxide and oxygen 总被引:27,自引:20,他引:7
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A number of methods were used to prepare a species of mammalian cytochrome oxidase (EC 1.9.3.1, ferrocytochrome c-oxygen oxidoreductase) in which only cytochrome a(3) is reduced and in combination with CO. The kinetics of CO binding by cytochrome a(3) (2+) in this species is significantly different from that exhibited by cytochrome a(3) (2+) in the fully reduced enzyme. The second-order rate constant for combination was 5x10(4)m(-1).s(-1) and the ;off' constant was 3x10(-2)s(-1). The kinetic difference spectra cytochrome a(3) (2+)-cytochrome a(3) (2+)-CO reveal further differences between the mixed-valence and the fully reduced enzyme. The reaction between cytochrome a(3) (2+) and oxygen in the mixed-valence species was followed in flow-flash experiments and reveals a fast, oxygen-dependent (8x10(7)m(-1).s(-1) at low oxygen) rate followed by a slow process, whose rate is independent of oxygen but whose amplitude is dependent on [O(2)]. The fast oxygen-dependent reaction yields as the first product the so-called ;oxygenated' enzyme. We conclude from these experiments that the ligand-binding behaviour of cytochrome a(3) depends on the redox state of its partners, a fact which represents clear evidence for site-site interaction in this enzyme. The fact that oxygen reacts rapidly with this enzyme species in which only one component, namely cytochrome a(3), is reduced represents clear and unequivocal evidence that this is indeed the O(2)-binding site in cytochrome oxidase and may indicate that reduction of oxygen can proceed via single electron steps. 相似文献
43.
Isolation and structure of nocobactin NA, a lipid-soluble iron-binding compound from Nocardia asteroides 总被引:3,自引:1,他引:2
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Nocobactin NA, a lipid-soluble iron-chelating product with an unusual and characteristic u.v.-absorption spectrum, was isolated from Nocardia asteroides grown under conditions of iron deficiency. Its structure was determined by physical methods and by synthesis of one of its degradation products. Nocobactin NA was obtained as a homologous mixture of compounds with side chains of differing length, and resembles mycobactin M in structure except that it has an oxazole ring in place of an oxazoline ring, and the side chains in the cobactin fragment are considerably shorter. 相似文献
44.
Stereospecific hydroxylation of 3-deoxy-1,2:5,6-di-O-isopropylidene-3-C-trans-and 3-C-cis-(methoxycarbonylmethylene)-α-D-ribo-hexofuranose (2 and 3, respectively), with potassium permanganate in pyridine afforded 3-C-[S- and R-hydroxy-(methoxycarbonyl)methyl]-1,2:5,6-di-O-isopropylidene-α-D-glucofuranose, (6 and 7, respectively), in a combined yield, after chromatography, of 43%. Selective formation of monomethanesulfonates (9a and 10a) and p-toluenesulfonates (9b and 10b), followed by treatment with sodium azide and reduction of the azide, afforded the methyl 2-D-(and 2-L-)(3-deoxy-1,2:5,6-di-O-isopropylidene-α-D-glucofuranos-3-yl)-glycinates (12a and 13a, respectively). Basic hydrolysis of the latter compounds yielded 2-D- and 2-L-(3-deoxy-1,2:5,6-di-O-isopropylidene-α-D-glucofuranos-3-yl)glycine (12b and 13b, respectively). The structures of the glycosyl amino acids were correlated with that of L-alanine by circular dichroism. 相似文献
45.
Investigation of Reported Aflatoxin Production by Fungi Outside the Aspergillus flavus Group 总被引:3,自引:2,他引:1
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Benjamin J. Wilson T. Colin Campbell A. Wallace Hayes Richard T. Hanlin 《Applied microbiology》1968,16(6):819-821
A screening study of 121 fungus isolates, representing 29 species, for aflatoxin synthesis demonstrated this property only in Aspergillus flavus and A. parasiticus. Eight of the organisms found negative were isolates reported by other investigators to produce aflatoxin. Since similar negative reports have come from several other workers, it is concluded that only the A. flavus group of Aspergillus can presently be certified as sources of these toxins. Reasons for possible false-positive findings are discussed along with precautionary measures and differential analytical procedures useful in aflatoxin screening studies. 相似文献
46.
Colin Ogilvie 《BMJ (Clinical research ed.)》1966,1(5489):735-736
47.
A method for the determination of desferrioxamine-available iron in tissue fractions is described which involves incubation with desferrioxamine, extraction of desferrioxamine and its iron-bound form, ferrioxamine, and quantitation of these two forms of the drug by reversed-phase hplc analysis. Chelatable iron levels in the 1-10µMolar region could be accurately and reproducibly measured using this technique.
The desferrioxamine-available iron levels in both the cortex and medulla of rabbit kidneys were significantly elevated (up to 2-fold) after the organs had been subjected to 2 hours warm ischaemia or 24 hours cold storage at 0°C In hypertonic citrate solution. There was no change in the total iron content of the tissues under these circumstances and thus a redistribution of intracellular iron to more available pools had presumably taken place as a result of ischaemia. This redistribution of iron may be an important factor in the initiation of peroxidative damage to cell membranes upon reperfusion of the organ with oxygen. 相似文献
The desferrioxamine-available iron levels in both the cortex and medulla of rabbit kidneys were significantly elevated (up to 2-fold) after the organs had been subjected to 2 hours warm ischaemia or 24 hours cold storage at 0°C In hypertonic citrate solution. There was no change in the total iron content of the tissues under these circumstances and thus a redistribution of intracellular iron to more available pools had presumably taken place as a result of ischaemia. This redistribution of iron may be an important factor in the initiation of peroxidative damage to cell membranes upon reperfusion of the organ with oxygen. 相似文献
48.
Dr. Bernard R. Glick Barbara J. Butler Colin I. Mayfield J. J. Pasternak 《Current microbiology》1989,19(3):143-146
We previously observed that whenAzotobacter vinelandii was transformed by different broad-host-range plasmids, normal cellular functions such as growth and siderophore production are impaired. In the present work, whenA. vinelandii was transformed with the low copy number plasmid pRK290, the extent of this metabolic impairment was lessened, as evidenced by increased siderophore production and moderate levels of growth on medium that lacks added iron. It is concluded that the severity of the plasmid-induced metabolic load reflects the relative level of expression of plasmid-encoded proteins. 相似文献
49.
The effect of dicyclohexylcarbodiimide (DCCD) on electron transfer in the acceptor quinone complex of reaction centers (RC) from Rhodobacter sphaeroides is reported. DCCD covalently labelled the RC over a wide concentration range. At low concentrations (<10 M) the binding was specific for the L subunit. At relatively high concentrations (>100 M) DCCD accelerated the rate of charge recombination of the P+QB
- state, consistent with a decrease in the equilibrium constant between QA
-QB and QAQB
-. At similar concentrations, in the presence of cytochrome c as exogenous donor, turnover of the RC was inhibited such that only three cytochromes were oxidized in a train of flashes. Both these inhibitory effects were fully reversed by dialysis, indicating that stable covalent binding was not involved. Possible mechanisms of action are discussed in terms of the putative role of specific residues in proton transfer and protonation and release of quinol from the RC. 相似文献
50.
Quinones may be toxic by a number of mechanisms. including arylation and oxidative stress caused by redox cycling. Using isolated hepatocytes, we have studied the cytotoxicity of four quinones. with differing abilities to arylate cellular nucleophiles and redox cycle. in relation to their effects on cellular pyridine nucleotides. High concentrations of menadione (redox cycles and arylates). 2-hydroxy-1,4-naphthoquinone (neither arylates nor redox cycles via a one electron reduction) 2.3-dimethoxy-1.4-naphthoquinone (a pure redox cycler) and p-benzoquinone (a pure arylator) caused an initial decrease in NAD+ and loss of viability, which was not prevented by 3-aminobenzamide. an inhibitor of poly(ADP-ribose)polymerase. In contrast. 3-aminobenzamide inhibited the loss of NAD' and viability caused by dimethyl sulphate so implicating poly(ADP-ribose)polymerase in its toxicity but not that of the quinones. Non-toxic concentrations of menadione. 2.3-dimethoxy-1.4-naphthoquinone and 2-hydroxy-1.4-naphthoquinone all caused markedly similar changes in cellular pyridine nucleotides. An initial decrease in NAD+ was accompanied by a small. transient increase in NADP+ and followed by a larger. prolonged increase in NADPH and total NADP+ + NADPH. Nucleotide changes were not observed with non-toxic concentrations of p-benzoquinone. Our findings suggest that a primary event in the response of the cell to redox cycling quinones is to bring about an interconversion of pyridine nucleotides. in an attempt to combat the effects of oxidative stress 相似文献