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1.
A difference in the incorporation of 14C into hippurate glycine from dl-[2-14C]glutamate and dl-[5-14C]glutamate in guinea pigs
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The specific radioactivity of urinary hippurate glycine was determined after injecting guinea pigs with benzoate and either dl-[2-(14)C]glutamate or dl-[5-(14)C]glutamate. The isotope dilution factor for the formation of [(14)C]glycine was significantly greater (30%) with C-2 labelled glutamate. With either form of labelled glutamate the hippurate glycine was largely carboxyl-group labelled. The observations suggest a route for the incorporation of glutamate carbon into glycine that involves C-5 but not C-2. A hypothesis for glycine biosynthesis from l-glutamate is advanced, consistent with these findings, that includes conversion of l-glutamate to 4-hydroxy-2-oxoglutarate, the scission of the latter to glyoxylate and pyruvate, and the formation of glycine by transamination. 相似文献
2.
The bacterial degradation of flavonoids. Oxidative fission of the A-ring of dihydrogossypetin by a Pseudomonas sp 总被引:2,自引:0,他引:2
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Cell-free extracts prepared from a Pseudomonas sp., grown on (+)-catechin, oxidized dihydrogossypetin (3',4',5,7,8-pentahydroxyflavanonol) by cleaving the A-ring to form oxaloacetic acid from C-5, C-6, C-7 and C-8 together with 5-(3,4-dihydroxyphenyl)-4-hydroxy-3-oxovalero-delta-lactone. The structure of this lactone was confirmed by synthesis of related phenylvalerolactones. 相似文献
3.
Yukiko Makino Shigeru Ohta Yoshikazu Tasaki Osamu Tachikawa Masaki Kashiwasake Masaaki Hirobe 《Journal of neurochemistry》1990,55(3):963-970
Serotonin binding protein (SBP) is a vesicular protein found in neurectoderm-derived cells that store 5-hydroxytryptamine (5-HT, serotonin), such as central and peripheral serotonergic neurons and paraneurons (parafollicular cells of the thyroid). 5-HT is stored as a complex with SBP in vivo. Two forms of the protein are found. These differ in molecular mass: one is 45 kDa and the other 56 kDa. It has been suggested that the 56-kDa form of SBP may be the precursor of the 45-kDa form. To study the relationship between these two proteins, we have used a covalently bound radiolabeled probe to analyze their binding domains. A photoaffinity reagent, N-(4-azido-2-nitrophenyl)-5-hydroxytryptamine (NAP-5-HT), was synthesized and characterized by nuclear magnetic resonance spectroscopy, mass spectra, and UV-visible absorption spectra. A 1 M excess of NAP-5-HT inhibited the binding of [3H]5-HT to SBP by 50%. NAP[3H]5-HT was also synthesized and attached to both high- and low-affinity binding sites on both forms of SBP. The high-affinity constants for 45-kDa and 56-kDa proteins were 0.8 nM and 0.02 nM, respectively, whereas the low-affinity constants were 0.3 microM and 0.15 microM. When the high-affinity site of partially purified SBP was photoaffinity-labeled with the reagent, two covalently labeled proteins (45 kDa and 56 kDa) were found by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Inhibition of the labeling of both proteins by 50% was observed in the presence of a 15-fold molar excess of 5-HT.(ABSTRACT TRUNCATED AT 250 WORDS) 相似文献
4.
The interactions of norharman (9H-pyrido [3,4-b] indole) and harman (1-methyl-9H-pyrido [3,4-b] indole) with DNA were studied. DNA caused remarkable fluorescence quenching and change in the absorption spectra of the dyes. Scatchard plots obtained by optical titration gave Kd values of 2.2 X 10(-5)M and 7.7 X 10(-6)M, and apparent numbers of binding sites of 0.13/base and 0.12/base for norharman and harman, respectively. Agarose gel electrophoresis of circular DNA, closed in the presence or absence of norharman revealed that the dye intercalates DNA, thereby causing 17 +/- 3 degrees unwinding of the double helix. 相似文献
5.
An a-mating-type-specific substance responsible for sexual agglutination was purified to 397-times in specific activity (units/mg protein) from the cytoplasm of a-mating type cells. The purified substance gave a single band stained with PAS reagent but not with both Coomassie brilliant blue and silver staining reagent by polyacrylamide gel electrophoresis in the presence of 8 M urea. However, incorporation of [35S]methionine and Lowry reaction clearly indicate that the substance is a glycoprotein. The substance specifically masked sexual agglutinability of cells of the opposite mating type , indicating univalent action. The substance is a glycoprotein with a carbohydrate content of 90%, a pI of 4.5, and a molecular weight of 130,000. The substance was inactivated by 2-mercaptoethanol and proteolytic enzymes but not by glycolytic enzymes. The substance formed a complementary complex having no biological activity when mixed with -agglutination substance from the wall or cytoplasm of -cells in vitro.Non-common abbreviations PAGE
polyacrylamide gel electrophoresis
- PAS
periodic acid-Schiff
- PBS
10-2 M phosphate buffer solution, pH 5.5
- PMSF
phenylmethyl sulfonyl fluoride
- SDS
sodium dodecyl sulfate 相似文献
6.
N A Markovich T I Vorotyntseva M I Zil'berman V K Antonov 《Biokhimii?a (Moscow, Russia)》1985,50(5):795-803
Cathepsin D was purified from the lactating rabbit mammary gland by a rapid procedure, which included fractionation with (NH4)2SO4, acid precipitation, double affinity chromatography on pepstatin-Sepharose 4B and gel filtration on Sephadex G-100, resulting in approximately 360-fold purification of the enzyme over the homogenate and approximately 16% recovery. After isoelectric focusing, the enzyme dissociated into four (pI 5.8, 6.3, 6.5 and 7.2) multiple forms, but appeared homogeneous on polyacrylamide gel electrophoresis. Cathepsin D has a Mr of 45 kDa as determined by Sephadex G-100 column chromatography. On sodium dodecylsulfate/polyacrylamide gel electrophoresis the enzyme gave a single protein band, corresponding to Mr of 45 kDa. The amino acid composition of the enzyme is similar to that of cathepsins D from other tissues. A single N-terminal amino acid was glycine. Cathepsin D contains 6.4% carbohydrates consisting of mannose, galactose, fucose and glucosamine at a ratio of 3:9:2:2. Cathepsin D is inhibited by pepstatin with Ki of 2.5 X 10(-9) M and irreversibly by N-diazoacetyl-N'-2.4-dinitrophenyl-ethylene diamine. The enzyme hydrolyzes bovine hemoglobin with the maximal activity at pH 3.0 with Km = 10(-5) M and HLeu-Ser-Phe(NO2)-Nle-Ala-Leu-OMe with Km = 4 X 10(-5) M and Rcat = 0.95 s-1. The major cleavage sites were Leu15-Tyr16, Phe24-Phe25 and Phe25-Tyr26 during hydrolysis of the oxidized insulin B-chain by cathepsin D. 相似文献
7.
Christina Bohlin Per Persson Lo Gorton Knut Lundquist Leif J. Jnsson 《Journal of Molecular Catalysis .B, Enzymatic》2005,35(4-6):100-107
The erythro form of the lignin model compound 1-(3,4-dimethoxyphenyl)-2-(2-methoxyphenoxy)-1,3-propanediol (1) was oxidized with laccase/ABTS, lead(IV) tetraacetate (LTA), lignin peroxidase/H2O2, cerium(IV) ammonium nitrate (CAN) and Fenton's reagent. The product profiles obtained with the different oxidants were compared after separation, identification and quantification of the products using HPLC, UV-diode array detector and electrospray ionization mass spectrometry in positive ionization mode. The oxidants generated different product profiles that reflected their different properties. Oxidation with laccase/ABTS resulted almost exclusively in formation of 1-(3,4-dimethoxyphenyl)-3-hydroxy-2-(2-methoxyphenoxy)-1-propanone (2). Oxidation with LTA resulted in more 3,4-dimethoxybenzaldehyde (3) than ketone 2. Lignin peroxidase and CAN gave similar product profiles and aldehyde 3 was the predominant product (only small amounts of ketone 2 were formed). Oxidation with Fenton's reagent resulted in the formation of more aldehyde 3 than ketone 2 but the yields were very low. CAN served as an excellent model for the lignin peroxidase-catalyzed oxidation, while the laccase-mediator system, LTA and Fenton's reagent provided distinctly different product profiles. Erythro-1-(3,4-dimethoxyphenyl)-1,2,3-propanetriol was present among the products obtained on oxidation with LTA, lignin peroxidase, CAN and Fenton's reagent. The differences in redox potential between the oxidants afford an explanation of the diverse product patterns but other factors may also be of importance. The reactions leading to cleavage of the β-ether bond with formation of 1-(3,4-dimethoxyphenyl)-1,2,3-propanetriol (veratrylglycerol) were found to proceed without affecting the configuration at the β-carbon atom. 相似文献
8.
Paul Scherer Vera Höllriegel Christine Krug Michael Bokel Paul Renz 《Archives of microbiology》1984,138(4):354-359
Exogenous 5-hydroxy-[2-14C]benzimidazole was transformed by Methanosarcina barkeri into 5-hydroxy-[2-14C]benzimidazolylcobamide. Thereby the endogenous biosynthesis of 5-hydroxybenzimidazole was completely blocked.Benzimidazole and 5,6-dimethylbenzimidazole were used by M. barkeri to form benzimidazolylcobamide respectively 5,6-dimethylbenzimidazolylcobamide (vitamin B12), but in these cases the endogenous biosynthesis of factor III was not completely suppressed.With [2-14C]benzimidazole it was demonstrated that this base as well as the benzimidazolylcobamide formed thereof are no precursors in the biosynthesis of 5-hydroxybenzimidazolylcobamide.Glycine instead was found to be a building block for the biosynthesis of 5-hydroxybenzimidazole, since radioactivity from [1-14C] and [2-14C]glycine was incorporated, into the base moiety of factor III, but not into its corrin moiety. With [1-13C]glycine and 13C-NMR-spectroscopy it was shown that C-1 of glycine gets C-3a of 5-hydroxybenzimidazole.[1-13C]glycine also led to a single prominent signal in the 13C-NMR-spectrum of coenzyme F420, this was assigned to C-10a.Thus C-1 of glycine was incorporated into the hydroxybenzene part of 5-hydroxybenzimidazole, whereas it was not incorporated into this part of coenzyme F420, indicating that the hydroxybenzene part of these two compounds is not formed from a common intermediate.
L-[U-14C]glutamate led to the exclusive labeling of the corrin ring of factor III, showing that the corrin precursor 5-aminolevulinic acid is formed by the C-5 pathway in M. barkeri.These experiments indicate that the biosynthesis of factor III in the archaebacterium M. barkeri is similar to the corrinoid biosynthesis in the anaerobic eubacteria Eubacterium limosum, Clostridium barkeri, and Clostridium thermoaceticum. 相似文献
9.
A kinetic analysis of [3H]-5-HT uptake in the photocytes of the photophores of Porichthys notatus revealed a high affinity (Km: 1.71 X 10(-7] and low affinity component (Km: 1.10 X 10(-5) M). The high affinity uptake was sodium- and potassium-dependent but largely insensitive to temperatures between 0 and 20 C. Ouabain (5 X 10(-3) M) and dinitrophenol (10(-3) M) reduced uptake significantly. DMI, imipramine and fluoxetine, in that order of potency, greatly inhibited [3H]-5-HT uptake. Noradrenaline and adrenaline reduced uptake in a non-competitive manner, while dopamine, tryptophan, 5-hydroxytryptophan and Cypridina luciferin had little or not effect on uptake. Non-facilitated luminescent responses to electrical stimulation were accompanied by release of [3H]-5-HT accumulated in the photocytes. Facilitatory luminescence excitation consistently failed to induce the release of [3H]-5-HT. Electrical and adrenaline (10(-5) M) stimulation of photophores after [3H]-5-HT release has occurred, failed to elicit any additional luminescent response. The photophores were responsive to KCN (10(-3) M) under these conditions. The results indicate that a specific carrier-mediated transport system is responsible for photocytic [3H]-5-HT uptake, and that release of photocytic [3H]-5-HT is stringently regulated and followed by inhibition of luminescence excitability. 相似文献
10.
Biosynthesis of thiamine. Incorporation experiments with 14C-labelled substrates and with [15N]glycine in Saccharomyces cerevisiae 总被引:4,自引:1,他引:3
1. Yeast was grown in a minimal synthetic medium together with a range of (14)C-labelled substrates under standardized conditions. After isolation, the purified thiamine was cleaved by sulphite and the pyrimidine and thiazole moieties were purified and assayed for radioactivity. 2. In order of decreasing incorporation, [(14)C]formate, [3-(14)C]serine, [2-(14)C]glycine and [2-(14)C]acetate supplied label for the pyrimidine, and [2-(14)C]glycine, [3-(14)C]serine, [1-(14)C]glycine, [(14)C]formate and [2-(14)C]acetate for the thiazole. Incorporation of label into the fragments from several other (14)C-labelled substrates, including [Me-(14)C]- and [3,4-(14)C(2)]-methionine, was insignificant. 3. [3-(14)C]Serine was shown not to contribute label to C-2 of the thiazole ring. 4. Significant incorporation of nitrogen from [(15)N]glycine into the thiazole moiety, but not into the pyrimidine moiety, was established. 5. It appears that C-2 and N-3 of the thiazole ring are formed from C-2 and the nitrogen atom of glycine, but the entire methionine molecule does not appear to be implicated. 相似文献
11.
Production of 3-Methyl-6-Methoxy-8-Hydroxy-3,4-Dihydroisocoumarin by Aspergillus caespitosus
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Aspergillus caepitosus Raper and Thom cultured on autoclaved cracked, yellow field corn produced 3-methyl-6-methoxy-8-hydroxy-3,4-dihydroisocoumarin which was extracted and then purified by silica gel columns. The physical and spectral properties of the colorless crystals from methylene chloride:hexane were consistent with those published for 3-methyl-6-methoxy-8-hydroxy-3,4-dihydroisocoumarin. Yield of high purity 3-methyl-6-methoxy-8-hydroxy-3,4-dihydroisocoumarin was about 140 mg of corn per kg. 相似文献
12.
We investigated the effect of octreotide (OCT), a stable somatostatin analog, (OCT) on changes in short-circuit current (Isc) induced by vasoactive intestinal peptide (VIP), aminophylline, serotonin (5-HT) and substance P. OCT significantly decreased basal Isc at a concentration of 10(-9) M; the maximum decrease in Isc was observed at 10(-6) M. OCT (10(-7) M) significantly inhibited the intestinal secretory response to all the secretagogues studied. The maximum Isc response was reduced when tissues were stimulated with VIP (184.9 +/- 18.0 vs. 119.7 +/- 14.1, P less than 0.05), 5-HT (135.1 +/- 14.4 vs. 79.5 +/- 13.4, P less than 0.05) and substance P (156.0 +/- 19.2 vs. 30.7 +/- 5.4, P less than 0.01). In the case of aminophylline, the concentration-response curve was shifted to the right but the maximum response was not reduced. Because VIP and aminophylline increase cAMP while 5-HT and substance P stimulate intestinal secretion principally by a calcium linked mechanism, we conclude that OCT inhibits Isc in rat colon by more than one mechanism. 相似文献
13.
Trevor Sharp Steven R. Bramwell David Clark David G. Grahame-Smith 《Journal of neurochemistry》1989,53(1):234-240
The effect of manipulating the activity of central 5-hydroxytryptamine (5-HT) neurones on extracellular 5-HT in ventral hippocampus of the chloral hydrate-anaesthetized rat was studied using the brain perfusion method, microdialysis. Basal levels of 5-HT in the dialysates were close to the detection limits of our assay using HPLC with electrochemical detection. However, addition of the selective 5-HT reuptake inhibitor citalopram (10(-6) M) to the perfusion medium produced readily measurable amounts of dialysate 5-HT. Citalopram, therefore, was used throughout our experiments. Hippocampal dialysate levels of 5-HT sharply declined over the first hour after dialysis probe implantation, but then became constant. This stable output of 5-HT was reduced by 57% in rats treated 14 days previously with intracerebroventricular injections of the 5-HT neurotoxin 5,7-dihydroxytryptamine. Electrical stimulation (1-ms pulse width, 300 microA, 2-20 Hz) of the dorsal raphe nucleus for 20 min caused a rapid rise in hippocampal 5-HT output, which immediately declined on cessation of the stimulus and was frequency-dependent. Addition of tetrodotoxin (10(-6) M) to the perfusion medium reduced 5-HT levels to 75% of predrug values. Injection of the 5-HT1A agonist 8-hydroxy-2-(di-n-propylamino)tetralin (0.5 and 2.5 micrograms) into the dorsal raphe nucleus caused a dose-related fall in hippocampal output of 5-HT compared to saline-injected controls. We conclude from these data that the spontaneous output of endogenous 5-HT into hippocampal dialysates, measured under our experimental conditions, predominantly originates from central 5-HT neurones and changes in accordance with their electrical activity. 相似文献
14.
Effect of serotonin on murine macrophages: suppression of Ia expression by serotonin and its reversal by 5-HT2 serotonergic receptor antagonists 总被引:2,自引:0,他引:2
Serotonin (5-HT), a mediator released from platelets at sites of inflammation, suppressed IFN-gamma-induced Ia expression in mouse bone marrow macrophages maintained in vitro. (Mean percent suppression = 63.9% +/- 9.2, n = 40.) This suppression was not toxic or endotoxin-related, was concentration-dependent, and occurred at the physiologic concentrations of 5-HT present at inflammatory sites. The concentration of 5-HT producing the half-maximal effect was 2.5 to 5.5 X 10(-8) M. Related compounds, dopamine, histamine, and tryptamine, were much less potent in suppressing IFN-gamma-induced Ia, with maximally suppressing concentrations more than 100-fold higher than the maximally suppressing 5-HT concentration. L-5-hydroxytryptophan (5-HTP), the most potent analog tested, was 10-fold less potent than 5-HT in suppressing Ia expression. The concentration of 5-HTP producing the half-maximal effect = 4 X 10(-7) M. 5-HT suppression of IFN-gamma-induced Ia expression was antagonized by the 5-HT2 type receptor antagonists spiperone, ketanserin, and LY53857. Concentrations of these agents resulting in 50% inhibition of the serotonin effect were 1.5 X 10(-8) M, 7.5 X 10(-8) M, and 4.5 X 10(-12) M, respectively. 5-HT was most effective in suppressing IFN-gamma-induced Ia when added early in culture simultaneously with IFN-gamma. These data provide functional evidence that 5-HT suppression of IFN-gamma-induced Ia expression is mediated through a 5-HT receptor with some characteristics of the 5-HT2 type. 5-HT may play a physiologic role at sites of inflammation as a modulator of the effects of IFN-gamma on macrophage function. 相似文献
15.
Chromatographic Analyses of the Serotonin 5-HT1A Receptor Solubilized from the Rat Hippocampus 总被引:1,自引:1,他引:0
S. El Mestikawy D. Taussig H. Gozlan M. B. Emerit M. Ponchant M. Hamon 《Journal of neurochemistry》1989,53(5):1555-1566
Serotonin 5-HT1A receptors in rat hippocampal membranes were solubilized by 10 mM 3-[3-(cholamidopropyl)dimethylammonio]-1-propane sulfonate (CHAPS) and chromatographed on various gels in an attempt to design a relevant protocol for their (partial) purification. In particular, an affinity gel made of the 8-hydroxy-2-(di-n-propylamino)tetralin (8-OH-DPAT) derivative 8-methoxy-2-[(N-propyl, N-butylamino)amino]tetralin (8-MeO-N-PBAT) coupled to Affigel 202 was specially developed for this purpose. First, studies of the effects of various compounds (detergents, lipids, reducing agents, sugars, etc.) on the specific binding of [3H]8-OH-DPAT and on the rate of heat-induced inactivation of solubilized 5-HT1A sites led to a buffer composed of 50 mM Tris-HCl, 50 microM dithiothreitol, 1 mM CHAPS, 10% glycerol, 0.1 mM MnCl2, and 50 micrograms/ml of cholesteryl hemisuccinate, pH 7.4, ensuring a high degree of stability of solubilized 5-HT1A sites, compatible with chromatographic analyses for 2-4 days at 4 degrees C. Adsorption and subsequent elution of [3H]8-OH-DPAT specific binding sites were found with several chromatographic gels, including wheat germ agglutinin-agarose, phenyl-Sepharose, hydroxylapatite-Ultrogel, diethylaminoethyl (DEAE)-Sepharose, and DEAE-Sephacel. Similarly, 8-MeO-N-PBAT-Affigel 202 allowed the adsorption and subsequent elution (by 1 mM 5-HT) of active 5-HT1A binding sites solubilized from rat hippocampal membranes. The two-step chromatography using 8-MeO-N-PBAT-Affigel 202 followed by wheat germ agglutinin-agarose gave a fraction enriched (by at least 400-fold) in 5-HT1A sites. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of this partially purified fraction revealed a major protein band with Mr close to 60,000. 相似文献
16.
Alkylation at N-1 of the NADP+ adenine ring with 3,4-epoxybutanoic acid gave 1-(2-hydroxy-3-carboxypropyl)-NADP+. Enzymic reduction of the latter, followed by alkaline Dimroth rearrangement and enzymic reoxidation, gave N6-(2-hydroxy-3-carboxypropyl)-NADP+. On the other hand, bromination at C-8 of the NADP+ adenine ring, followed by reaction with the disodium salt of 3-mercaptroproionic acid, gave 8-(2-carboxyethylthio)-NADP+. Carbodimide coupling of the three carboxylic NADP+ derivatives to polyethyleneimine afforded the corresponding macromolecular NADP+ analogues. The carboxylic and the polyethyleneimine derivatives synthesized have been shown to be co-enzymically active with yeast glucose-6-phosphate dehydrogenase, liver glutamate dehydrogenase and yeast aldehyde dehydrogenase. The degree of efficiency relative to NADP+ with the three enzymes ranged from 17% to 100% for the carboxylic derivatives and from 1% to 36% for the polyethyleneimine analogues. On comparing the efficiences with the three enzymes of the N-1 derivatives to the one of the corresponding N6 anc C-8 analogues, the order of activity was N-1 greater than N6 greater C-8, except in the case of the carboxylic compounds with glutamate dehydrogenase, where this order was inverted. None of these modified cofactors were active with pig heart isocitrate dehydrogenase. 相似文献
17.
Guanine deaminase (EC 3.5.4.3, guanine aminohydrolase [GAH]) was purified 3248-fold from human liver to homogeneity with a specific activity of 21.5. A combination of ammonium sulfate fractionation, and DEAE-cellulose, hydroxylapatite, and affinity chromatography with guanine triphosphate ligand were used to purify the enzyme. The enzyme was a dimer protein of a molecular weight of 120,000 with each subunit of 59,000 as determined by gel filtration and sodium dodecyl sulfate-gel electrophoresis. Isoelectric focusing gave a pI of 4.76. It was found to be an acidic protein, as evidenced by the amino acid analysis, enriched with glutamate, aspartate, alanine and glycine. It showed a sharp pH optimum of 8.0. The apparent Km for guanine was determined to be 1.53 X 10(-5) M at pH 6.0 and 2 X 10(-4) M for 8-azaguanine as a substrate at pH 6.0. The enzyme was found to be sensitive to p-hydroxymercuribenzoate inhibition with a Ki of 1.53 X 10(-5) M and a Ki of 5 X 10(-5) M with 5-aminoimidazole-4-carboxamide as an inhibitor. The inhibition with iodoacetic acid showed only a 7% loss in the activity at 1 X 10(-4) M and a 24% loss at 1 X 10(-3) M after 30 min of incubation, whereas p-hydroxymercuribenzoate incubation for 30 min resulted in a 91% loss of activity at a concentration of 1 X 10(-4) M. Guanine was the substrate for all of the inhibition studies. The enzyme was observed to be stable up to 40 degrees C, with a loss of almost all activity at 65 degrees C with 30 min incubation. Two pKa values were obtained at 5.85 and 8.0. Analysis of the N-terminal amino acid proved to be valine while the C-terminal residue was identified as alanine. 相似文献
18.
D-Glucose (0.7 M), glycine (0.3 M), and sodium hydrogencarbonate (0.1 M) were dissolved in aqueous 30% ethanol at pH 8.0 and left at 50 °C for 4 d in a dark room under nitrogen displacement. The resulting blue pigment was isolated and purified from the blue solution by anionic exchange and gel filtration chromatography. This blue pigment, which is designated Blue-G1, was identified as 5-[1,4-bis-carboxymethyl-5-(2,3,4-trihydroxybutyl)-1,4-dihydropyrrolo[3,2-b]pyrrol-2-ylmethylene]-1,4-bis-carboxymethyl-2-(2,3,4-trihydroxybutyl)-4,5-dihydropyrrolo[3,2-b]pyrrol-1-ium. Blue-G1 had two symmetrical pyrrolopyrrole structures with a trihydroxybutyl group. Blue-G1 had a polymerizing activity, suggesting it to be an important Maillard reaction intermediate through the formation of melanoidins. 相似文献
19.
Rat liver tyrosine aminotransferase was purified by chromatography on CM-Sephadex C-50 and DEAE-cellulose, (NH4)2SO4 fractionation and gel filtration on Sephadex G-200. Livers from 400 rats can be easily worked up by this procedure. Furthermore, this purification method has the advantage that hepatic tryptophan 2,3-dioxygenase, which, like tyrosine aminotransferase, is induced by glucocorticosteroids, can be purified from the same homogenate. Tyrosine aminotransferase purified by this method was shown to be specific for 2-oxoglutarate. Its subunits have a molecular weight of 45 000. The following "apparent" Michaelis constants were determined: L-tyrosine, 1.7 X 10(-3) M; 2-oxoglutarate, 5.9 X 10(-4) M; and pyridoxal 5'-phosphate, 2.1 X 10(-6) M. Tyrosine aminotransferase, depleted of its cofactors, binds 4 molecules of pyridoxal 5'-phosphate per 90 000 daltons with a KA of 2.2 X 10(5) M-1. 相似文献
20.
5-Hydroxytryptamine (serotonin or 5-HT) stimulated the incorporation of 32Pi into phosphatidylinositol (PI) but not into polyphosphoinositides in C6 glioma cells with an EC50 of 1.2 X 10(-7) M. The phosphoinositide response was blocked by the 5-HT2 antagonists ketanserin and spiperone but inhibited only partly by methysergide and mianserin. Atropine, prazosin, and yohimbine did not block the response, whereas fluphenazine and haloperidol did so partially but also inhibited basal incorporation by approximately 30%. The 5-HT1A agonist 8-hydroxy-2(di-n-propylamino)tetralin did not cause stimulation. Incubation with 5-HT (1 microM) for 1 h increased the incorporation of [2-3H]myoinositol into all phosphoinositides but not into inositol phosphates (IPs). Li+ alone at 10 mM increased labeling in inositol bisphosphate (IP2) and trisphosphate (IP3), whereas labeling in IP and phosphoinositides remained unaltered. Addition of 5-HT had no effect on this increase. Mn2+ at 1 mM enhanced labeling in PI, PI-4-phosphate, lyso-PI, glycerophosphoinositol, and IP, but the presence of 5-HT again did not cause further stimulation. 5-HT also stimulated the release of IPs in cells prelabeled with [2-3H]myo-inositol, incubated with LiCl (10 mM) and inositol (10 mM), and then exposed to 5-HT (1 microM). Radioactivity in IP2 and IP3 was very low, was stimulated approximately 50% as early as 30 s, and remained elevated for at least 20 min. Radioactivity in IP was at least 10 times as high as in IP3 but was increased only from 3 min on with a peak at 20 min, when the elevation was approximately 40 times that in IP3.(ABSTRACT TRUNCATED AT 250 WORDS) 相似文献