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141.
Activation of wheat chloroplast sedoheptulose bisphosphatase: a continuous spectrophotometric assay 总被引:3,自引:0,他引:3
A new continuous spectrophotometric assay for sedoheptulose 1,7-bisphosphatase, applied to studies of the activation and steady-state kinetics of the wheat enzyme, is described. The assay enzyme sequence couples the formation of sedoheptulose 7-phosphate to the oxidation of NADH. The recycling of the reaction substrate enables measurements to be made at essentially constant substrate concentrations. Activation of wheat chloroplast sedoheptulose 1,7-bisphosphatase required a reducing agent and could be described by a first-order rate constant. The rate of activation was greatly increased in the presence of Mg2+ and sedoheptulose 1,7-bisphosphate. The Km of the activated enzyme for sedoheptulose 1,7-bisphosphate. and its S0.5 for Mg2+ were found to be 13.3 μm and 1.6 mm respectively. A high recovery method for purifying wheat chloroplast sedoheptulose 1,7-bisphosphatase is also detailed. 相似文献
142.
In rat liver membranes cholera toxin ADP-ribosylated two polypeptides (Mr 42000 and 47000) in the regulatory component of adenylate cyclase. L-arginine methyl ester specifically inhibited both the activation of adenylate cyclase and ADP-ribosylation by cholera toxin, suggesting that cholera toxin modified arginine, or arginine-like, residues. A hydrolysis-resistant analogue of GTP (β, γ-imidoguanosine 5′-triphosphate, p(NH)ppG) bound to the regulatory protein in an essentially irreversible manner. Pretreatment with the analogue failed to inhibit the labelling of polypeptides by cholera toxin showing that the sites for ADP-ribosylation were different from those at which guanyl nucleotides were bound. 相似文献
143.
Cytoplasmic proteases of rat liver parenchymal cells 总被引:2,自引:0,他引:2
G N DeMartino 《Biochemical and biophysical research communications》1982,108(3):1325-1330
Soluble extracts of isolated rat liver parenchymal cells contained three proteases with alkaline pH optima. One protease was a high molecular weight (Mr = 500,000) enzyme which was stimulated by ATP. The other two proteases were totally dependent on calcium for activity and displayed different calcium concentration requirements. One was half-maximally activated by 150 μM Ca2+ while the other required only 10 μM Ca2+ for half-maximal activation. 相似文献
144.
E Melloni B Sparatore F Salamino M Michetti S Pontremoli 《Biochemical and biophysical research communications》1982,107(3):1053-1059
The soluble neutral proteinase of human erythrocytes dissociates into constituent subunits of 80k and 30k in the presence of mM concentrations of Ca2+. Similarly the soluble natural inhibitor of this proteinase, of approximate molecular weight 240k, is dissociated into 60k subunits by mM concentrations of Ca2+. Removal of Ca2+ restores the native oligomeric structure of the proteinase and of the natural inhibitor. The formation of the native active enzyme or of the inactive enzyme-inhibitor complex depends on reversible association-dissociation processes mediated by Ca2+ concentration. 相似文献
145.
Isolated, intact rat liver mitochondria, without extraneous substrates but loaded with Ca2+ (20 nmol/mg), can be observed to release Ca2+ when treated with ruthenium red. Such release can be inhibited by 0.33 mM dlisocitrate but not by 10 mM dl-β-hydroxybutyrate. Assays of NADP+, NADPH, NAD+, and NADH revealed that only the reduction of NADP+ can be linked with such inhibition of Ca2+ release, not that of NAD+. Since ruthenium redinsensitive Ca2+ release is a physiological (but normally masked) process, this experimental approach avoids some potential problems ascribed to strong pyridine nucleotide oxidation. It is suggested that specific NADP+:NADPH dependent reactions are part of a physiological mechanism regulating Ca2+ release/retention. 相似文献
146.
Induction and repair of DNA strand breaks in cultured human fibroblasts exposed to various phenols and dihydrodiols of benzo[a]pyrene 总被引:1,自引:0,他引:1
Cultured human fibroblasts from healthy donors were incubated for 30 min with nine different benzo[a]pyrene (BP) derivatives in the presence or absence of liver microsomes from 3-methylcholanthrene treated rats. The induction and repair of DNA strand breaks were analysed by alkaline unwinding and separation of double and single stranded DNA (SS-DNA) by hydroxylapatite chromatography immediately after the incubation or at various times after the treatment. In the absence of microsomes DNA stand breaks were detected in fibroblasts exposed to 30 microM of each of the six BP phenols (1-, 2-, 3-, 7-, 9- or 11-OH-BP) and the three BP dihydrodiols (BP-4,5-, BP-7,8- or BP-9,10-dihydrodiol). After removal of the BP derivatives from the medium the DNA strand breaks disappeared within 24 h. alpha-Naphthoflavone (alpha-NF) caused a decrease in the induction of strand breaks by 1-, 3- and 9-OH-BP but did not affect the induction of strand breaks in cells exposed to BP-7,8-dihydrodiol. In the presence of microsomes DNA strand breaks were found after exposure to 30 microM of each of the six BP phenols (1-, 2-, 3-, 7-, 9- or 11-OH-BP), as well as BP-7,8- and 9,10-dihydrodiol. In contrast BP-4,5-dihydrodiol did not induce strand breaks under these conditions. The induction of strand breaks by BP-7,8-dihydrodiol was enhanced in the presence of cytosine-1-beta-D-arabinofuranoside (AraC). In all cases the DNA strand breaks had disappeared 24 h after removal of the BP derivatives and microsomes except after treatment with BP-7,8-dihydrodiol. 相似文献
147.
Takeaki Miyamoto Hiroshi Sakabe Hiroshi Inagaki 《International journal of biological macromolecules》1983,5(3):188-190
The β-structure of S-caboxymethyl derivatives of microfibrillar proteins isolated from Merino wool was investigated by X-ray diffraction for comparison with the structur of β-keratin. The S-carboxymethylated microfibrillar proteins(SCMKA) w well-oriented β-films of SCMKA weer obtained by stretching the SCMKA cast films in steam up to about 300% extesnsion. It was found that the reflections in β-pattern of SCMKA may be indexed on a pseudo-orthorhombic unit cell with a =0.94 nm, b = 0.66 nm and c = nm, where the ab, and c axes are in the direction of the interchain hydrogen bonding, the main chain(fibre axis) and the side chain, respectively. The unit cell dimesnions evaluated for SCMKA were almost the same as those for β-keratin, suggeting that few peptide sequences containing S-carboxymethyl cystine may be involved in the formation of β-structure from SCMKA. 相似文献
148.
The objective was to determine the effect of ACTH 1-17, an adrenocorticotropin analogue, on the mitotic index in the corneal epithelium of mice standardized in 12 hr of light alternating with 12 hr darkness. A question asked was whether the time of administration along the 24-hr time scale influenced any response found. The findings showed that ACTH 1-17 could, depending upon when it was administered, bring about a statistically significant decrease, an increase or even no such change in the mitotic index. The greatest responses found were increases, especially when ACTH 1-17 was administered during the dark span. Also the time after injection when the responses occurred varied. The greatest response recorded was at 12 hr after injection when ACTH 1-17 was given at 2 hr into the dark with a 641% and a 718% increase with a low (0.02 IU/kg) and a higher (20 IU/kg) dose, respectively. A 3-way analysis of variance supported the conclusion that the kind-of-treatment, time-of-treatment and treatment-to-kill interval (sampling time) are important factors when determining any response to ACTH 1-17 on the mitotic index. 相似文献
149.
Mhamed Sa?d El Kebbaj Norbert Latruffe Yves Gaudemer 《Biochemical and biophysical research communications》1982,108(1):42-50
In order to titrate and understand the role of arginyl residues of D-β-hydroxybutyrate dehydrogenase, arginyl specific reagents: butanedione, 1,2-cyclohexanedione and phenylglyoxal were incubated with three different forms of the enzyme; native enzyme (inner mitochondrial membrane bound), purified apoenzyme (phospholipid -free) and phospholipid-enzyme complex (reconstituted active form).After complete inactivation of the enzyme by [14C]-phenylglyoxal, the number of modified arginyl residues was different: one with the lipid-free apoenzyme and three with the phospholipid-enzyme complex, suggesting a conformational change of the enzyme triggered by the presence of phospholipids.After exhaustive chemical modification either of the apoenzyme or of the phospholipid-enzyme complex with [14C]-phenylglyoxal, four arginyl residues were titrated indicating that these residues are located in the hydrophilic part of the enzyme, not interacting with phospholipids.Reconstituted enzyme inactivated by butanedione could no longer bind a pseudosubstrate (succinate) which indicates that an arginyl residue is involved in the enzyme-substrate complex formation.The values of second order rate constants of D-β-hydroxybutyrate dehydrogenase inactivation by butanedione and 1,2-cyclohexanedione were unchanged with the three enzyme forms, suggesting that phospholipids are not involved in the substrate binding mechanism. 相似文献
150.