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Photodynamic effects were demonstrated and assayed under field conditions in a number of different laboratory strains and pond isolates of cyanobacteria; parameters assayed for resistance to photooxidation were viable count, turbidity of the cyanobacterial suspension, and protein and pigment contents. The effects of density, colonial structure, and internal gas vacuoles on the lethal outcome were investigated. The stability and formation of superoxide dismutase under photooxidative conditions in the field and laboratory were studied in the different strains. An isolate of Microcystis from blooms in ponds exhibited extremely high resistance to photooxidation, which was abolished by exposure to chloramphenicol. 相似文献
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In order to investigate the nature of amino acid residues involved in the active in the active site of a ribonuclease from Aspergillus saitoi, the pH dependence of the rates of inactivation of RNase Ms by photooxidation and modification with diethylpyrocarbonate were studied. (1) RNase Ms was inactivated by illumination in the presence of methylene blue at various pH's. The pH dependence of the rate of photooxidative inactivation of RNase Ms indicated that at least one functional group having pKa 7.2 was involved in the active site. (2) Amino acid analyses of photooxidized RNase Ms at various stages of photooxidative inactivation at pH's 4.0 and 6.0 indicated that one histidine residue was related to the activity of RNase Ms, but that no tryptophan residue was involved in the active site. (3) 2',(3')-AMP prevented the photooxidative inactivation of RNase Ms. The results also indicated the presence of a histidine residue in the active site. (4) Modification of RNase Ms with diethylpyrocarbonate was studied at various pH's. The results indicated that a functional group having pKa 7.1 was involved in the active site of RNase Ms. 相似文献
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Partial inactivation of tau-dinitrophenylhistidine-200 human carbonic anhydrase B, induced by visible light, followed first order kinetics (k(app) = 6.05 times 10-2 min-1). After 50 min the tau-dinitrophenylhistidine (tau-DNP-histidine) content decreased to a negligible level, but the illuminated enzyme retained, at pH 7.6, approximately 9.2 percent of the esterase activity of the native enzyme. The following lines of evidence suggest that the loss of activity results from the destruction of tau-DNP-histidine-200. (1) No significant loss of amino acid other than tau-DNP-histidine was detected after illumination. (2) The rate of loss of activity correlated well with the loss of tau-DNP-histidine. (3) In the photooxidized enzyme the DNP moiety was retained but had lost the characteristic sensitivity of tau-DNP-histidine to nucleophilic attack. Titration of the illuminated enzyme with acetazolamide indicated that the residual activity is an intrinsic property of the modified enzyme. The chromatographically purified photooxidized enzyme migrated as a single band on isoelectrofocusing in polyacylamide gel, and at pH 7.6 possessed 7.5 percent esterase activity relative to the native enzyme. By establishing effective destruction of histidine-200, it can be concluded that neither the pi N nor, as previously shown, the tau N of histidine-200 is critical for the catalysis. 相似文献
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Irradiation with visible light of human serum albumin in aqueous solution at pH 8, in the presence of catalytic amounts of rose bengal or methylene blue, resulted in random oxidation of the histidine residues in the protein under consumption of one mole O2, and release of somewhat less than one proton, per histidine residue degraded. An increase of light absorption at 250 nm was proportional to the amount of oxygen consumed. Bilirubin bound to the oxidized protein showed an increased light absorption at its maximum, 460 nm, and a decreased binding affinity, indicating a conformational change of the protein on oxidation of histidine residues. This change also resulted in a slight perturbation of tyrosine light absorption, corresponding to a shift of the chromophore to more polar surroundings. Further, a sensitized oligomerization of albumin was observed, independent of oxidation of the histidine residues, and not consuming oxygen. Irradiation of a complex of human serum albumin with one molecule of bound bilirubin, in the absence of a sensitizing dye, resulted in a fast, non-oxygen consuming process whereby the light absorption maximum of the pigment was shifted 4 nm towards longer wavelength and part of the bilirubin was converted to a more polar pigment, bound less firmly to the protein. This was followed by a relatively slow oxidation of the pigment under uptake of one mole O2. Parallel photooxidation of the protein carrier could not be detected. It is considered possible that the fast, anaerobic process is operative in phototherapy of hyperbilirubinemia in the newborn. Serum albumin is probably not oxidized during this treatment. 相似文献
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Photooxidation of alpha-glucan phosphorylases from rabbit muscle and potato tubers in the presence of rose bengal leads to a rapid loss of enzymatic activity which follows first-order kinetics. The process is pH dependent, being more rapid at higher pH. The inactivation is closely related to the destruction of histidine residues in the enzyme. It is suggested that histidine residues are largely responsible for the loss of enzymatic activity in the photooxidation. The inactivation of potato phosphorylase is retarded by substrates, whereas that of the muscle enzyme is not. The rate of photoinactivation of muscle phosphorylase b is increased with AMP, and decreased with ATP, ADP, IMP and glucose-6-P. This finding is considered to be closely related to the allosteric transition of phosphorylase. 相似文献
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Methylene blue immobilized on porous glass beads was used to catalyze the photooxidation of methionine alone and the methionine residues of lysozyme. A solution of 2 mM methionine in 50% acetic acid was oxidized to methionine sulfoxide in the presence of immobilized methylene blue after 6 h of photooxidation at 37 degrees C. Selective photooxidation of the methionyl residues in lysozyme was achieved after 26 h of reaction in 84% acetic acid at 4 degrees C. The specific activity of lysozyme exposed to light in the presence of methylene blue decreased by 94%, while that of a lysozyme solution in the presence of methylene blue not exposed to light decreased by 21%. The lysozyme solution exposed to light but not containing the methylene blue beads lost 33% of its specific activity after the same period of photooxidation. It was shown that the decrease in enzyme activity was not caused by adsorption of the enzyme onto the beads. 相似文献
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Cytochrome b559 (cyt b559) is an intrinsic and essential component of the photosystem II (PSII) protein complex, but its function, stoichiometry, and electron-transfer kinetics in the physiological system are not well-defined. In this study, we have used flash-detection optical spectroscopy to measure the kinetics and yields of photooxidation and dark reduction of cyt b559 in untreated, O2-evolving PSII-enriched membranes at room temperature. The dark redox states of cyt b559 and the primary electron acceptor, QA, were determined over the pH range 5.0-8.5. Both the fraction of dark-oxidized cyt b559 and dark-reduced QA increased with increasing acidity. Consistent with these results, an acid-induced drop in pH from 8.5 to 4.9 in a dark-adapted sample caused the oxidation of cyt b559, indicating a shift in the redox state during the dark reequilibration. As expected from the dark redox state of cyt b559, the rate and extent of photooxidation of cyt b559 during continuous illumination decreased toward more acidic pH values. After a single, saturating flash, the rate of photooxidation of cyt b559 was of the same order of magnitude as the rate of S2QA- charge recombination. In untreated PSII samples at pH 8.0 with 42% of cyt b559 oxidized and 15% of QA reduced in the dark, 4.7% of one copy of cyt b559 was photooxidized after one flash with a t1/2 of 540 +/- 90 ms. On the basis of our previous work [Buser, C. A., Thompson, L. K., Diner, B. A., & Brudvig, G. W (1990) Biochemistry 29, 8977] and the data presented here, we conclude that Sn+1, YZ., and P680+ are in redox equilibrium and cyt b559 (and YD) are oxidized via P680+. After a period of illumination sufficient to fully reduce the plastoquinone pool, we also observed the pH-dependent dark reduction of photooxidized cyt b559, where the rate of reduction decreased with decreasing pH and was not observed at pH < 6.4. To determine the direct source of reductant to oxidized cyt b559, we studied the dark reduction of cyt b559 and the reduction of the PQ pool as a function of 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU) concentration. We find that DCMU inhibits the reduction of cyt b559 under conditions where the plastoquinone pool and QA are reduced. We conclude that QB-. (H+) or QBH2 is the most likely source of the electron required for the reduction of oxidized cyt b559.(ABSTRACT TRUNCATED AT 400 WORDS) 相似文献
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The principal products of the photooxidation of 7,12-dimethylbenz[a]-anthracene (DMBA) in aqueous solutions by photooxidation induced by laboratory lighting have been characterized by high performance liquid chromatograms (HPLC), ultraviolet and mass spectrograms and by comparisons with authentic samples. The products identified were the 7,12-epidioxy-7,12-dihydro-7-12-dimethyl-, 7,12-dione, 7-hydroxymethyl-12-methyl-, 12-hydroxymethyl-7-methyl-, 7-formyl-12-methyl-, 12-formyl-7-methyl-, and 12-hydroxy-12-methyl-7-one derivatives of benz[a]-anthracene. The HPLC profile of products is similar to that obtained from oxidation of DMBA by 'one-electron' reagents, singlet oxygen, or liver microsomal metabolism. The first points of attack are the 7- and 12- positions. The mechanism of photooxidation appears to be generation of singlet oxygen by photodynamic effect of DMBA. None of the products is photosensitizing, however. 相似文献
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Photooxidation of primaquine (1) and 5-hydroxyprimaquine (5) afforded a blue dye for which o-quinone structure 4 was elaborated. Similar oxidation of N-ethoxyacetylprimaquine (10) afforded o-quinone 11. Tissue schizontocidal activity of 4 and 11, and bisquinolylmethine 3 prepared earlier, showed that none of them had noteworthy antimalarial activity, but all three produced methemoglobin. 相似文献
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Photooxidation of the protoporphyrin-apomyoglobin complex 总被引:2,自引:0,他引:2
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Subtilisins Novo and DY were photoinactivated in the presence of methylene blue according to first order kinetics. The competitive inhibitor N alpha-benzoyl-L-arginine protected significantly against inactivation. Under the conditions employed in this study a selective photooxidation of the active site histidine 64 was achieved. Rate constants of 0.32 X 10(-2), s-1 and 0.35 X 10(-2), s-1, were calculated for the Novo enzyme and subtilisin DY, respectively. Apparent pKa values of the catalytically important imidazole group of 7.0 +/- 0.1 (s. Novo) and 7.1 +/- 0.1 (s. DY) were directly determined. The histidyl residues in the two proteases, except the active site histidine, which is the first target of photooxidation, are "buried" in the interior of the protein globule. Conformational studies suggested that the photoreactive histidine is not involved in the stabilization of the protein conformation. 相似文献
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J Stuart I N Pessah T G Favero J J Abramson 《Archives of biochemistry and biophysics》1992,292(2):512-521
The photooxidizing xanthene dye rose bengal is shown to induce rapid Ca2+ release from skeletal muscle sarcoplasmic reticulum (SR) vesicles. In the presence of light, nanomolar concentrations of rose bengal increase the Ca2+ permeability of the SR and stimulate the production of singlet oxygen (1O2). In the absence of light, no 1O2 production is measured. Under these conditions, higher concentrations of rose bengal (micromolar) are required to stimulate Ca2+ release. Furthermore, removal of oxygen from the release medium results in marked inhibition of the light-dependent reaction rate. Rose bengal-induced Ca2+ release is relatively insensitive to Mg2+. At nanomolar concentrations, rose bengal inhibits [3H]ryanodine binding to its receptor. beta,gamma-Methyleneadenosine 5'-triphosphate, a nonhydrolyzable analog of ATP, inhibits rose bengal-induced Ca2+ release and prevents rose bengal inhibition of [3H]ryanodine binding. Ethoxyformic anhydride, a histidine modifying reagent, at millimolar concentrations induces Ca2+ release from SR vesicles in a manner similar to that of rose bengal. The molecular mechanism underlying rose bengal modification of the Ca2+ release system of the SR appears to involve a modification of a histidyl residue associated with the Ca2+ release protein from SR. The light-dependent reaction appears to be mediated by singlet oxygen. 相似文献
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Ryo Yamauchi Masahiro Kojima Masatoshi Isogai Koji Kato Yoshimitsu Ueno 《Bioscience, biotechnology, and biochemistry》2013,77(11):2815-2820
Spinach monogalactosyldiacylglycerol (MGDG) and digalactosyldiacylglycerol (DGDG) were oxidized with singlet molecular oxygen by the use of chlorophyll a as the photosensitizer. The oxidation products were separated from the unoxidized MGDG and DGDG by reverse-phase high performance liquid chromatography (HPLC). The products separated by HPLC were identified to be mono- and di-hydroperoxides formed by 1O2 oxidation of the 16:3 or 18:3 component of MGDG and DGDG. Each unsaturated fatty acid moiety in the MGDG and DGDG produced isomeric hydroperoxides in a manner similar to the corresponding fatty acid methyl ester. 相似文献
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Light dependent oxygen metabolism of chloroplast preparations. III. Photooxidation of ascorbic acid 总被引:1,自引:1,他引:0
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Helen M. Habermann 《Plant physiology》1961,36(2):252-261
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14CO2 uptake in leaves of wheat plants (Triticum aestivum L.) fertilized by urea or Ca(NO3)2 (25 mol m-3) was investigated. The Warburg effect (inhibition of 14CO2 uptake by oxygen) under 0.03 vol. % CO2 concentration was observed only in non-fertilized plants. Under 0.03 vol. % CO2, the Warburg antieffect (stimulation of 14CO2 uptake by oxygen) was detected only in plants fertilized by Ca(NO3)2. Under saturating CO2 concentration (0.30 vol. %), the Warburg antieffect was observed in all variants. Under limitation of ribulose-1,5-bisphosphate carboxylase/oxygenase activity (0.30 vol. % CO2 + 1 vol. % O2), the rate of synthesis of glycollate metabolism products decreased in control and urea-fertilized plants but was enhanced in nitrate-fed plants. Hence, there was an activation of glycollate formation via transketolase reaction in fertilized plants, and the products of nitrate reduction function were oxidants in nitrate-fertilized plants whereas the superoxide radical played this role in urea-fertilized plants. 相似文献
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Papain [EC 3.4.22.2] was photooxidized using methylene blue as a sensitizer. The photooxidzed enzyme lost its caseinolytic activity and had significantly decreased histidine and tryptophan contents. The tyrosine content was the same before and after the photooxidation. The SH content of the photooxidized enzyme, as determined after reduction with dithiothreitol, was also unchanged. The loss of histidine was always slower than the loss of enzymatic activity, being less than one residue per molecule even when the enzymatic activity was completely lost. However, the inactivation and the oxidation of a histidine residue were pH-dependent in a similar fashion in the pH range of 5.0-8.0, the pH profiles conforming to theoretical titration curves with apparent pKa values of 6.6 and 6.7, respectively. The fact that the ionization of a histidine residue in papain has a normal imidazole pKa value is entirely in accord with the finding for stem bromelain [EC 3.4.22.4] (Murachi, T., Tsudzuki, T., & Okumura, K. (1975) Biochemistry 14, 249-255), and is of great significance in relation to the mechanism of catalysis by these enzymes. 相似文献
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