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A flavoenzyme which showed NADPH-cytochrome c reductase (NADPH-cytochrome c oxidoreductase EC 1.6.2.4) and transhydrogenase (NADPH-NAD+ oxidoreductase, EC 1.6.1.1) activities was purified to an electrophoretically homogeneous state from Nitrobacter winogradskyi. The reductase was a flavoprotein which contained one FAD per molecule but no FMN. The oxidized form of the enzyme showed absorption maxima at 272, 375 and 459 nm with a shoulder at 490 nm, its molecular weight was estimated to be 36,000 by SDS polyacrylamide gel electrophoresis, and the enzyme seemed to exist as a dimer in aqueous solution. The enzyme catalyzed reduction of cytochrome c, DCIP and benzylviologen by NADPH, oxidation of NADPH with menadione and duroquinone, and showed transhydrogenase activity. NADH was less effective than NADPH as the electron donor in the reactions catalyzed by the enzyme. The NADPH-reduction catalyzed by the enzyme of N. winogradskyi cytochrome c-550 and horse cytochrome c was stimulated by spinach ferredoxin. The enzyme reduced NADP+ with reduced spinach ferredoxin and benzylviologen radical.Abbreviations DCIP
dichlorophenolindophenol
- Tris
trishydroxy-methylaminomethane
- Mops
3-(N-morpholino) propanesulfonic acid
- SDS
sodium dodecylsufate 相似文献
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Transformation of mouse BALB/c 3T3 cells with human basic fibroblast growth factor cDNA. 总被引:22,自引:3,他引:19 下载免费PDF全文
The expression of human basic fibroblast growth factor (bFGF) cDNA in mouse BALB/c 3T3 clone A31 cells induced morphological transformation. These transformed cells grew well and reached more than a sixfold-higher saturation density than parental A31 cells even in serum-free medium. They were able to form colonies in soft agar. The phenotypic alteration in the transformed cells was reversed by the addition of anti-human bFGF antibodies to the medium. These results suggest that the cellular transformation mediated by bFGF is caused by autocrine stimulation with secreted bFGF molecules. 相似文献
5.
Makoto Kurokawa Kiyoaki Kuwasawa 《Journal of comparative physiology. A, Neuroethology, sensory, neural, and behavioral physiology》1988,162(4):533-541
1. | The excitatory and inhibitory influences on the gill ofAplysia Juliana, which are mediated by the branchial nerve, were studied by means of electrophysiological techniques. Excitatory and inhibitory pathways in the nerve were stimulated simultaneously or selectively. |
2. | The branchial nerve was found to contain both excitatory and inhibitory pathways which did not contain synapses in the branchial ganglion. The excitatory pathways caused longitudinal shortening of the gill along the efferent branchial vessel and the inhibitory pathways were modulatory, depressing the longitudinal shortening. |
3. | Branchial nerve stimulation elicited two types of excitatory junctional potential (EJP), which were not mediated by the branchial ganglion, in a muscle cell of the efferent branchial vessel. One type was attributed to the central motor neuron and the other type to a motor neuron which is probably situated in the neural plexus of the gill periphery. |
4. | Four inhibitory pathways from the central nervous system to the gill were found. |
5. | Inhibitory junctional potentials (IJPs) recorded from muscle cells of the efferent branchial vessel in response to branchial nerve stimulation did not have monosynaptic characteristics. It is thought that inhibitory motor neurons which were activated by the branchial nerve might exist at the neural plexus of the gill. |
6. | A single EJP which has been induced by a stimulus pulse applied to the excitatory pathway of the branchial nerve may be depressed in an all-or-none manner by a stimulus pulse applied to the inhibitory pathway, if this is done within a distinct short period prior to or after the stimulus inducing the EJP. This indicates that the central motor neuron receives presynaptic inhibition at its periphery. |
7. | The motor neurons of the neural plexus seem to receive inhibitory innervation. Suppression of endogenous EJPs in the efferent vessel persisted for a long period even after cessation of stimulation. |
8. | A certain branchioganglionic neuron (BGN) was found to receive inhibitory postsynaptic potential (IPSP) inputs from the branchial nerve. |
9. | The multimodality of both the excitatory and the inhibitory pathways in the branchial nerve may explain the compound neural modulations of gill movements. |
6.
Hangil Chang Naohide Yamashita Hiroshi Matsunaga Kiyoshi Kurokawa 《The Journal of membrane biology》1988,103(3):263-271
Summary We have previously reported hyperpolarizing membrane potential changes in a monkey kidney cell line (JTC-12) which has characteristics resembling proximal tubular cells. These hyperpolarizations could be observed spontaneously or evoked by mechanically touching adjacent cells. In this report, we have shown further evidence that these hyperpolarizations are elicited by an increase in membrane conductance to K+ which is caused by an increase in cytosolic Ca2+ concentration. In addition, we have found another type of hyperpolarization which is evoked by applying flow of extracellular fluid to the cell. Intracellular injection of Ca2+ and Sr2+ evoked hyperpolarizations, while intracellular injection of Mn2+ and Ba2+ did not. Intracellular injection of EGTA suppressed both spontaneous and mechanically evoked hyperpolarizations. In Ca2+-free medium, both spontaneous and flow-evoked hyperpolarizations were not observed, while mechanical stimuli consistently evoked hyperpolarization. In Na+-free medium, the incidence of cells showing the spontaneous or flow-evoked hyperpolarization increased, and the amplitude and the duration of the mechanically evoked hyperpolarization became greater. Quinidine inhibited all types of hyperpolarization. These data suggest that hyperpolarizations in JTC-12 cells are due to an increase in Ca2+-activated K+ conductance. 相似文献
7.
Inhibition of rat brain adenylate cyclase activity by benzodiazepine through the effects on Gi and catalytic proteins 总被引:1,自引:0,他引:1
The effect of benzodiazepines on adenylate cyclase system was examined in rat brain. Micromolar concentrations of diazepam inhibited the enzyme activity in synaptic membranes in dose- and time-dependent manners. The inhibitory effect of diazepam was more evident on the enzyme activity in the presence of guanylyl-5'-imidodiphosphate (GppNHp) or NaF-AlCl3 than on that in the basal state. In the pertussis toxin-treated membranes, the effect of diazepam in the presence of GppNHp or NaF-AlCl3 was markedly suppressed. In addition, other benzodiazepines, such as medazepam, flurazepam, flunitrazepam, and clonazepam, had similar effects to those of diazepam, whereas Ro15-1788, an antagonist of a high affinity receptor in the central nervous system, had no effect on adenylate cyclase activity and did not antagonize the effect of diazepam. These findings indicate that benzodiazepines inhibit rat brain adenylate cyclase activity through the effects on both a low affinity benzodiazepine receptor coupled with the inhibitory GTP-binding regulatory protein (Gi) and catalytic protein. 相似文献
8.
M Yoneda K Takatsuki Y Oiso T Takano M Kurokawa A Ota A Tomita T Ohno K Okano T Kanazawa 《Endocrinologia japonica》1986,33(1):89-94
The serum bone Gla-protein (BGP) and urinary gamma-carboxyglutamic acid (gamma-Gla) levels were determined in patients with primary hyperparathyroidism (PHP). The mean serum BGP and urinary gamma-Gla levels were 18.6 +/- 2.34 ng/ml and 65.5 +/- 4.62 nmoles/mgCr, respectively, for the 11 patients with the skeletal type of PHP, 5.13 +/- 0.85 ng/ml and 45.2 +/- 1.33 nmoles/mgCr for the 4 with the chemical type, and 7.91 +/- 2.43 ng/ml and 43.2 +/- 3.47 nmoles/mgCr for the 5 with the renal type. Thus, patients with skeletal-type PHP had significantly higher serum BGP and urinary gamma-Gla levels than those with the other type of PHP. Serum BGP levels had significant positive correlations with serum Ca (r = 0.64, P less than 0.005), serum A1-p (r = 0.77, P less than 0.001) and serum PTH (r = 0.45, P less than 0.005). Urinary gamma-Gla levels also had significant positive correlations with serum Ca (r = 0.50, P less than 0.05), serum A1-p (r = 0.67, P less than 0.005), serum 1,25(OH)2D (r = 0.62, P less than 0.02), and serum BGP (r = 0.72, P less than 0.001). Mineral content in the left radius had significant negative correlations with serum BGP levels (r = -0.73, P less than 0.001) and urinary gamma-Gla levels (r = -0.59, P less than 0.01). As these data show, serum BGP and urinary gamma-Gla levels clearly reflect the abnormal bone metabolism and can therefore be useful biochemical markers in PHP. 相似文献
9.
Reverse-phase high-performance liquid chromatography (RP-HPLC) has been examined as an approach to the rapid analysis of carcinogen-modified histones. H1 and core histone fractions were prepared by differential acid extraction of 0.35 M NaCl-extracted rat liver nuclei previously exposed to [3H]-7r,8t-dihydroxy-9t, 10t-oxy-7,8,9, 10-tetrahydrobenzo(a)pyrene [( 3H]BPDE-I). Using a sodium perchlorate-phosphate (PCP)/acetonitrile solvent system, the H1 histone fraction was eluted from an Aquapore RP-300 column in five peaks (P1-P5). The core histone fraction was resolved into eight peaks (C1-C8) using a PCP/acetonitrile-methanol solvent system. The histones of each peak were identified by sodium dodecyl sulfate and Triton/acid/urea gel electrophoresis or amino acid analysis as follows: P1, H1 degrees; P2-P5, four different H1 variant fractions; C1, H4 + A24; C2, H2B; C3, H2A X 2 + to one H2A variant; C4, H2A.1; C5, H2A.1 + two H2A variants; C6, H3.2; C7, H3.3; C8, H3.1. The bulk of radioactivity was covalently bound to histone H2A, which had higher specific activities of BPDE-I than other histones. Significant amounts of radioactivity were observed in histones H3 and H1, but not in histones H2B and H4. These RP-HPLC systems have the advantages of an analysis time within 60 min, the identification of H1, H2A, and H3 variants, and the quantitative analysis of radioactive histones. These results indicate that these RP-HPLC systems are very useful to analyze the binding of carcinogens to histones. 相似文献
10.