RESUMEN
A large number of new glycans, derived from glycoproteins, has been characterized in the past few years. O-linked fucose was found in epidermal growth factor-like domains of several proteins. For the N-linked glycans of Helix pomatia hemocyanin, novel types of antennae were identified. The positions of noncarbohydrate substituents were established in N-glycans. C-mannosylation of a tryptophan residue was discovered in human ribonuclease 2 and is the first example of C-glycosylation in glycoproteins.
Asunto(s)
Endorribonucleasas/metabolismo , Glicosilación , Polisacáridos/metabolismo , Proteínas/metabolismo , Animales , Sitios de Unión , Secuencia de Carbohidratos , Humanos , Datos de Secuencia Molecular , Polisacáridos/química , Proteínas/química , SulfatosRESUMEN
Excitotoxicity is a paradigm used to explain the biochemical events in both acute neuronal damage and in slowly progressive, neurodegenerative diseases. Here, we show in a longitudinal magnetic resonance imaging study that Delta(9)-tetrahydrocannabinol (Delta(9)-THC), the main active compound in marijuana, reduces neuronal injury in neonatal rats injected intracerebrally with the Na(+)/K(+)-ATPase inhibitor ouabain to elicit excitotoxicity. In the acute phase Delta(9)-THC reduced the volume of cytotoxic edema by 22%. After 7 d, 36% less neuronal damage was observed in treated rats compared with control animals. Coadministration of the CB(1) cannabinoid receptor antagonist SR141716 prevented the neuroprotective actions of Delta(9)-THC, indicating that Delta(9)-THC afforded protection to neurons via the CB(1) receptor. In Delta(9)-THC-treated rats the volume of astrogliotic tissue was 36% smaller. The CB(1) receptor antagonist did not block this effect. These results provide evidence that the cannabinoid system can serve to protect the brain against neurodegeneration.
Asunto(s)
Edema Encefálico/prevención & control , Cannabis , Dronabinol/farmacología , Fármacos Neuroprotectores/farmacología , Ouabaína/toxicidad , Enfermedad Aguda , Animales , Animales Recién Nacidos , Encéfalo/efectos de los fármacos , Encéfalo/metabolismo , Encéfalo/patología , Edema Encefálico/inducido químicamente , Edema Encefálico/diagnóstico , Edema Encefálico/metabolismo , Enfermedad Crónica , Cuerpo Estriado/efectos de los fármacos , Cuerpo Estriado/metabolismo , Cuerpo Estriado/patología , Relación Dosis-Respuesta a Droga , Inhibidores Enzimáticos/farmacología , Estudios Longitudinales , Imagen por Resonancia Magnética , Microinyecciones , Ouabaína/administración & dosificación , Ratas , Ratas Wistar , Receptores de Cannabinoides , Receptores de Droga/antagonistas & inhibidores , Receptores de Droga/metabolismo , ATPasa Intercambiadora de Sodio-Potasio/antagonistas & inhibidores , Agua/metabolismoRESUMEN
The endocannabinoid anandamide [N-arachidonoylethanolamine (AEA)] is thought to function as an endogenous protective factor of the brain against acute neuronal damage. However, this has never been tested in an in vivo model of acute brain injury. Here, we show in a longitudinal pharmacological magnetic resonance imaging study that exogenously administered AEA dose-dependently reduced neuronal damage in neonatal rats injected intracerebrally with the Na(+)/K(+)-ATPase inhibitor ouabain. At 15 min after injury, AEA (10 mg/kg) administered 30 min before ouabain injection reduced the volume of cytotoxic edema by 43 +/- 15% in a manner insensitive to the cannabinoid CB(1) receptor antagonist SR141716A. At 7 d after ouabain treatment, 64 +/- 24% less neuronal damage was observed in AEA-treated (10 mg/kg) rats compared with control animals. Coadministration of SR141716A prevented the neuroprotective actions of AEA at this end point. In addition, (1) no increase in AEA and 2-arachidonoylglycerol levels was detected at 2, 8, or 24 hr after ouabain injection; (2) application of SR141716A alone did not increase the lesion volume at days 0 and 7; and (3) the AEA-uptake inhibitor, VDM11, did not affect the lesion volume. These data indicate that there was no endogenous endocannabinoid tone controlling the acute neuronal damage induced by ouabain. Although our data seem to question a possible role of the endogenous cannabinoid system in establishing a brain defense system in our model, AEA may be used as a structural template to develop neuroprotective agents.
Asunto(s)
Ácidos Araquidónicos/farmacología , Lesiones Encefálicas/prevención & control , Neuronas/efectos de los fármacos , Animales , Animales Recién Nacidos , Western Blotting , Encéfalo/efectos de los fármacos , Encéfalo/metabolismo , Encéfalo/patología , Edema Encefálico/inducido químicamente , Edema Encefálico/patología , Edema Encefálico/prevención & control , Lesiones Encefálicas/inducido químicamente , Lesiones Encefálicas/patología , Moduladores de Receptores de Cannabinoides , Cannabinoides/metabolismo , Modelos Animales de Enfermedad , Relación Dosis-Respuesta a Droga , Endocannabinoides , Inhibidores Enzimáticos , Glicéridos/metabolismo , Estudios Longitudinales , Imagen por Resonancia Magnética , Microinyecciones , Neuronas/metabolismo , Neuronas/patología , Ouabaína , Piperidinas/farmacología , Alcamidas Poliinsaturadas , Pirazoles/farmacología , Ratas , Ratas Wistar , Receptores de Cannabinoides , Receptores de Droga/antagonistas & inhibidores , RimonabantRESUMEN
Type 1 vanilloid receptors (VR1) have been identified recently in the brain, in which they serve as yet primarily undetermined purposes. The endocannabinoid anandamide (AEA) and some of its oxidative metabolites are ligands for VR1, and AEA has been shown to afford protection against ouabain-induced in vivo excitotoxicity, in a manner that is only in part dependent on the type 1 cannabinoid (CB1) receptor. In the present study, we assessed whether VR1 is involved in neuroprotection by AEA and by arvanil, a hydrolysis-stable AEA analog that is a ligand for both VR1 and CB1. Furthermore, we assessed the putative involvement of lipoxygenase metabolites of AEA in conveying neuroprotection. Using HPLC and gas chromatography/mass spectroscopy, we demonstrated that rat brain and blood cells converted AEA into 12-hydroxy-N-arachidoylethanolamine (12-HAEA) and 15-hydroxy-N-arachidonoylethanolamine (15-HAEA) and that this conversion was blocked by addition of the lipoxygenase inhibitor nordihydroguaiaretic acid. Using magnetic resonance imaging we show the following: (1) pretreatment with the reduced 12-lipoxygenase metabolite of AEA, 12-HAEA, attenuated cytotoxic edema formation in a CB1 receptor-independent manner in the acute phase after intracranial injection of the Na+/K+-ATPase inhibitor ouabain; (2) the reduced 15-lipoxygenase metabolite, 15-HAEA, enhanced the neuroprotective effect of AEA in the acute phase; (3) modulation of VR1, as tested using arvanil, the VR1 agonist capsaicin, and the antagonist capsazepine, leads to neuroprotective effects in this model, and arvanil is a potent neuroprotectant, acting at both CB1 and VR1; and (4) the in vivo neuroprotective effects of AEA are mediated by CB1 but not by lipoxygenase metabolites or VR1.
Asunto(s)
Ácidos Araquidónicos/fisiología , Cannabinoides/farmacología , Capsaicina/análogos & derivados , Capsaicina/metabolismo , Ácidos Grasos Insaturados/fisiología , Lipooxigenasa/fisiología , Degeneración Nerviosa/prevención & control , Fármacos Neuroprotectores/farmacología , Receptores de Droga/fisiología , Animales , Animales Recién Nacidos , Células Sanguíneas/efectos de los fármacos , Células Sanguíneas/enzimología , Células Sanguíneas/metabolismo , Encéfalo/efectos de los fármacos , Encéfalo/enzimología , Encéfalo/metabolismo , Química Encefálica , Mapeo Encefálico , Moduladores de Receptores de Cannabinoides , Endocannabinoides , Etanolaminas/análisis , Etanolaminas/metabolismo , Lipooxigenasa/metabolismo , Masculino , Masoprocol/farmacología , Degeneración Nerviosa/inducido químicamente , Degeneración Nerviosa/enzimología , Ouabaína/farmacología , Alcamidas Poliinsaturadas , Ratas , Ratas Wistar , Receptores de Droga/metabolismoRESUMEN
Analysis of the sialic acids obtained by mild acid hydrolysis of B lymphocytes reveals the presence of N-acetylneuraminic acid and 9-O-acetyl-N-acetylneuraminic acid. For T lymphocytes only N-acetylneuraminic acid has been demonstrated to occur. The applied methods include quantitative colorimetry, thin-layer chromatography and combined gas-liquid chromatography-mass spectrometry.
Asunto(s)
Linfocitos B/metabolismo , Ácidos Siálicos/metabolismo , Linfocitos T/metabolismo , Cromatografía en Capa Delgada , Colorimetría , Cromatografía de Gases y Espectrometría de Masas , Humanos , Ácidos Siálicos/sangreRESUMEN
Antiserum directed against lipoxygenase-1 from soybean (Glycine Max (L). Merr. var. Williams) was developed in rabbits with electrophoretically pure lipoxygenase-1 as an antigen. To remove traces of lipoxygenase-1 from a lipoxygenase-2 preparation the latter was chromatographed over a column of Sepharose 4B to which antibodies directed against lipoxygenase-1 were coupled. During affinity chromatography of lipoxygenase-2 no protein and only a small amount of activity were lost. Affinity-purified enzyme was used for immunization of rabbits to produce lipoxygenase-2 antibodies. Results with double gel immunodiffusion tests with the two soybean lipoxygenases and antisera directed against them demonstrated that there is no immunological relationship between the isoenzymes. Lipoxygenases from different species of leguminosae crossreacted only with antiserum directed against soybean lipoxygenase-2, and not with anti soybean lipoxygenase-1 serum. No crossreaction could be detected between soybean lipoxygenase antisera and lipoxygenases from species of Gramineae, Linaceae and Solanaceae.
Asunto(s)
Sueros Inmunes , Lipooxigenasa/inmunología , Plantas/enzimología , Complejo Antígeno-Anticuerpo , Cromatografía de Afinidad , Inmunodifusión , Glycine maxRESUMEN
The ability of soybean lipoxygenases-1 and -2 to oxygenate biomembranes isolated from soybean seedlings has been investigated. Constituents of the lipid bilayer were analyzed by means of reversed phase and chiral phase high performance liquid chromatography, gas chromatography/mass spectrometry, high performance thin layer chromatography and uv spectroscopy. Evidence is presented that soybean lipoxygenase-2, at variance with the type-1 enzyme, oxygenates the esterified unsaturated fatty acid moieties in biomembranes, whereas membrane-embedded free unsaturated fatty acid moieties were not a suitable substrate for either isoenzyme. The oxygenation products derived from the biomembranes were the 9- and 13-hydroperoxides of linoleic acid residues, in a molar ratio of 1.0 to 1.7, and the 9- and 13-hydroperoxides of alpha-linolenic acid residues, in a molar ratio of 1.0 to 0.1. The R/S ratios of 13-hydroperoxy-9Z,11E-octadecadienoic acid and 9-hydroperoxy-10E,12Z,15Z-octadecatrienoic acid were found to be 0.5 and 25.0, respectively. These stereospecificity values were much higher than those of hydroperoxides isolated after incubation of lipoxygenase-2 with non-membraneous fatty acids or their methyl esters. The hydroperoxy fatty acids produced were distributed in neutral lipids and phospholipids isolated from soybean membranes, the former being oxidized to a larger extent. Furthermore, both intracellular and plasma membranes were substrates for the enzymic oxygenation, with a preference for those of chloroplasts followed by those of Golgi apparatus, endoplasmic reticulum, plasma membrane and mitochondria. These data point towards a different action of the two lipoxygenases in soybean cells. We suggest that the type-2 enzyme plays a role in the in vivo remodelling of biomembranes. The physiological relevance of these findings is discussed.
Asunto(s)
Glycine max/enzimología , Membrana Dobles de Lípidos/química , Lipooxigenasa/metabolismo , Oxígeno/análisis , Lipooxigenasa/aislamiento & purificación , Glycine max/química , Glycine max/ultraestructura , Fracciones Subcelulares/químicaRESUMEN
Protoplasts were isolated from lentil (Lens culinaris) roots and their suitability as a transient expression system was investigated. After transfecting the protoplasts with the beta-glucuronidase (GUS) gene by either electroporation or polyethylene glycol (PEG), the specific activity of the reporter enzyme and the cell viability were determined. Electroporation was more effective than PEG treatment as transfection procedure and its efficiency was affected by the plasmid length. The feasibility of electro-transferring at the same time (coelectroporation) inhibitory anti-lipoxygenase monoclonal antibodies and the GUS-carrying plasmid pBI 221 was investigated as well. The amount of transferred immunoglobulins was quantitated by ELISA and the inhibitory ability of monoclonal antibodies on the intracellular target enzyme was determined. Evidence is presented for the successful coelectroporation of immunoglobulins and plasmid DNA into lentil protoplasts, the two types of macromolecules acting independently of each other in the recipient cells.
Asunto(s)
Anticuerpos Monoclonales , Electroporación , Fabaceae/genética , Plantas Medicinales , Plásmidos/genética , Transfección/métodos , Fabaceae/citología , Expresión Génica , Genes Reporteros , Glucuronidasa/biosíntesis , Glucuronidasa/genética , Lipooxigenasa/genética , Raíces de Plantas/citología , Raíces de Plantas/genética , ProtoplastosRESUMEN
Fe(III)-soybean lipoxygenase-1 yields with 4-nitrocatechol a green coloured 1 : 1 complex, which shows at pH 7.0 absorption maxima at 385 nm and 650 nm. The formation of this complex is reversible. The circular dichroism spectrum of the complex of Fe(III)-lipoxygenase-1 and 4-nitrocatechol has a positive band at around 380 nm and a negative band at around 450 nm and is significantly different from that of the Fe(III)-enzyme as such. 4-Nitrocatechol can be displaced from the green complex by 13-L-hydroperoxy-cis-9, trans-11-octadecadienoic acid, resulting in the formation of the blue complex between the Fe(III)-enzyme and 13-L-hydroperoxy-cis-9,trans-11-octadecadienoic acid both under aerobic and anaerobic conditions. Also linoleic acid competes with 4-nitrocatechol for the binding site on the Fe(III)-enzyme, as can be demonstrated under anaerobic conditions, ultimately leading to reduction of the Fe(III)-enzyme. The oxygenation of linoleic acid by Fe(III)-lipoxygenase-1 is inhibited by 4-nitrocatechol. From steady-state kinetics a non-competitive inhibition pattern is obtained. Probably it has to be considered as pseudo non-competitive because of the slow establishment of the complex equilibrium. An inhibition constant (K4NC) of 16.3 microM is found. On prolonged incubation of Fe(III)-lipoxygenase-1 and 4-nitrocatechol the green complex converts into a brown species. This conversion is found to be coupled with a change in the nature of the inhibition from reversible to irreversible. A complex between native lipoxygenase-1 and 4-nitrocatechol is found to be unlikely.
Asunto(s)
Catecoles/análisis , Compuestos Férricos/análisis , Hierro/análisis , Lipooxigenasa/análisis , Plantas/enzimología , Dicroismo Circular , Compuestos Férricos/metabolismo , Cinética , Ácidos Linoleicos/metabolismo , Lipooxigenasa/metabolismo , Conformación Molecular , Nitrocompuestos/análisis , Nitrocompuestos/metabolismo , Glycine max , Análisis EspectralRESUMEN
Soybean lipoxygenase-1 was found to contain five free sulfhydryl groups and no disulfide bridges. Three sulfhydryl groups react readily with methylmercuric halides. This modification results in significant changes of the catalytic properties of the enzyme. Comparison of modified and native lipoxygenase-1 shows the following: 1. The catalytic constant of the oxygenation of linoleic acid is reduced by approximately 50%, whereas the affinity towards linoleic acid remains unaltered. 2. At high concentrations of substrate and low concentrations of enzyme the kinetic lag phase in the oxygenation is considerably longer. 3. The regio- and stereospecificities of the oxygenation are significantly lower. 4. Besides hydroperoxides, oxo-octadecadienoic acids (4%) are formed during the oxygenation. 5. The cooxidation capacity is considerably enhanced. Treatment of methylmercury-modified lipoxygenase-1 with NaHS results in the complete recovery of the sulfhydryl groups and of the catalytic properties.
Asunto(s)
Lipooxigenasa/metabolismo , Plantas/enzimología , Disulfuros , Cinética , Ácidos Linoleicos , Compuestos de Metilmercurio , Glycine max , Relación Estructura-Actividad , Especificidad por Sustrato , Compuestos de Sulfhidrilo , Sulfuros/farmacologíaRESUMEN
Crude immunoglobulin G (IgG) fractions of antisera directed against soybean lipoxygenase-1 and -2 were purified by being passed through an immunoadsorbent column containing lipoxygenase coupled to CNBr-activated Sepharose 4B. Bound immunoglobulin was desorbed with pulses of 2 M or 3 M ammonium thiocyanate or 0.1 M glycine-HCl buffer (pH 2.5). The total column recoveries of anti-lipoxygenase-1 IgG and anti-lipoxygenase-2 IgG were 45% and 58%, respectively. The affinity for lipoxygenase of immunospecific antibodies was determined in an enzyme-linked immunosorbent assay (ELISA). In a reaction with lipoxygenase-1, anti-lipoxygenase-1 IgG, which was eluted with glycine-HCl buffer (pH 2.5) with recovery of 24%, had a 6.5-times higher affinity than the whole IgG fraction of antiserum. The affinity of anti-lipoxygenase-2 IgG for lipoxygenase-2 increased 2.2-times after chromatography of IgG over an immunoadsorbent column using 2 M ammonium thiocyanate as eluent (recovery 21%).
Asunto(s)
Anticuerpos/aislamiento & purificación , Inmunoglobulina G/aislamiento & purificación , Lipooxigenasa/análisis , Plantas/enzimología , Complejo Antígeno-Anticuerpo , Cromatografía de Afinidad/métodos , Ensayo de Inmunoadsorción Enzimática , Glycine maxRESUMEN
The leukotriene production by bovine polymorphonuclear leukocytes isolated from peripheral blood has been studied. Cells were incubated in the presence of arachidonic acid, glutathione, calcium ionophore A23187 and Ca2+. The leukotrienes then formed are leukotriene C4, leukotriene B4, two all-trans isomers of leukotriene B4 and the double dioxygenation product 12-epi-6-trans-8-cis-leukotriene B4. Leukotriene C4 is formed in such a large quantity by the bovine polymorphonuclear leukocyte that it might constitute an excellent and inexpensive source for the biosynthetic preparation of this spasmogenic leukotriene.
Asunto(s)
Leucotrieno B4/biosíntesis , Neutrófilos/metabolismo , SRS-A/biosíntesis , Animales , Calcimicina/farmacología , Calcio/farmacología , Bovinos , Cinética , Leucotrieno B4/sangre , Neutrófilos/efectos de los fármacos , Neutrófilos/inmunología , SRS-A/sangreRESUMEN
The novel sialic acid 4-O-acetyl-9-O-lactyl-N-acetylneuraminic acid has been identified as a constituent of horse submandibular gland glycoproteins in addition to the already known equine sialic acids, N-acetylneuraminic acid, 4-O-acetyl-N-acetylneuraminic acid, 9-O-acetyl-N-acetylneuraminic acid, 4,9-di-O-acetyl-N-acetylneuraminic acid, N-glycolylneuraminic acid, 4-O-acetyl-N-glycolylneuraminic acid and 9-O-acetyl-N-glycolylneuraminic acid. The structure has been established by combined gas-liquid chromatography-mass spectrometry.
Asunto(s)
Ácidos Siálicos/aislamiento & purificación , Glándula Submandibular/análisis , Animales , Cromatografía de Gases y Espectrometría de Masas , CaballosRESUMEN
The interaction of nitric oxide with the non-heme iron dioxygenase lipoxygenase is reported. This apparently resulted in a novel type of complex where an electron is donated to the NO molecule. In addition a new position for an EPR transition from iron was discovered which, it is suggested results from high spin ferric iron in a field of axial symmetry characterised by a very low value for D.
Asunto(s)
Lipooxigenasa , Espectroscopía de Resonancia por Spin del Electrón , Hierro , Ligandos , Ácidos Linoleicos , Óxido NítricoRESUMEN
A method is presented for determination of the enantiomeric composition of hydroxyperoxides formed by enzymic oxygenation of unsaturated fatty acids. After reduction of the hydroperoxy group with NaBH4, and esterification, the positional isomers of the resulting hydroxy compounds are separated by high performance liquid chromatography. The latter are subsequently subjected to a chiral derivatization to form diastereomeric alpha-methoxy-alpha-trifluoromethylphenylacetate esters. Determination of the diastereomeric composition by a NMR shift experiment furnishes the enantiomeric composition of the parent hydroperoxides. The method has been applied to the hydroperoxides formed by incubation of linoleic acid by corn germ or soybean lipoxygenase. Our results indicate that under the conditions used the hydroperoxides are mainly enantiospecifically formed.
Asunto(s)
Peróxidos Lipídicos/análisis , Lipooxigenasa/metabolismo , Espectroscopía de Resonancia Magnética/métodos , Borohidruros , Ácidos Linoleicos , EstereoisomerismoRESUMEN
Type-2 lipoxygenases from soybeans and peas, which have a pH optimum of 6--7 were examined for oxygenation activity at pH 9.0. The reaction velocity was found to be strongly dependent on substrate concentration. At higher substrate concentrations an inhibitory effect was observed, which is connected with the occurrence of a kinetic lag phase. On incubation of linoleic acid at pH 9.0 with either of these enzymes predominantly 9-LR-hydroperoxy-10-trans,12-cis-octadecadienoic acid is formed. The similarity of the product specificity with that of prostaglandin synthetase is discussed in view of the formation of prostaglandin-like substances by soybean lipoxygenase-2 (Bild, G.S., Bhat, S.G., Ramadoss, C.S. and Axelrod, B. (1978) J. Biol. Chem, 253, 21--23).
Asunto(s)
Ácidos Linoleicos/metabolismo , Peróxidos Lipídicos/biosíntesis , Lipooxigenasa/metabolismo , Plantas/enzimología , Dicroismo Circular , Fabaceae/enzimología , Cinética , Ácidos Linoleicos/aislamiento & purificación , Plantas Medicinales , Glycine maxRESUMEN
Soybean lipoxygenase-1 reacts with both 9-D and 13-L-hydroperoxylinoleic acids under anaerobic conditions. Approximately 40% of the hydroperoxide is converted into oxodienes, absorbing at 285 nm. Concomitantly, more polar compounds are formed, tentatively identified as being mainly epoxy-hydroxy-octadecenoic acids. When oxygen is present, the reaction is strongly inhibited, until in a very slow reaction the oxygen has been depleted. This accounts for the occurrence of a lag period.
Asunto(s)
Glycine max , Ácidos Linoleicos/metabolismo , Lipooxigenasa/metabolismo , Anaerobiosis , Concentración de Iones de Hidrógeno , Consumo de Oxígeno , Peróxidos/metabolismoRESUMEN
Lipoxygenase-1 from soybeans is incubated with an isomer of linoleic acid, 13-cis, 16-cis-octadecadienoic acid. Analysis of the oxygenation products indicates that molecular oxygen is stereospecifically introduced mainly at C-17 (n-2) of the fatty acid (in the LS-configuration), and only to a minor extent at C-13 (n-6). These findings contradict previous suggestions about the postional specificity of lipoxygenase-1.
Asunto(s)
Ácidos Linoleicos/metabolismo , Lipooxigenasa/metabolismo , Isomerismo , Oxidación-Reducción , Glycine max , Relación Estructura-ActividadRESUMEN
The kinetic parameters of the first and second oxygenation of arachidonic acid by soybean lipoxygenase-1 were determined and found to be for the first step at pH 10.0, Km (arachidonic acid) = 8.5 +/- 0.5 microM; kcat = 225 +/- 7 s-1 and for the second step at pH 8.7 Km (15-HPETE) = 440 +/- microM; kcat = 25 +/- 1 s-1. In the second oxygenation for which 15-Ls-hydroperoxy 5-cis, 8-cis, 11-cis 13-trans-eicosatetraenoic acid is a substrate, two isomeric dihydroperoxy fatty acids are formed. After separation of the corresponding dihydroxy esters by high-performance liquid chromatography, they were identified by mass-spectrometry, 1H- and 13C-NMR spectroscopy as 8-DS, 15-LS-dihydroperoxy 5-cis, 9-trans, 11-cis, 13-trans-eicosatetraenoic acid and 5-DS, 15-LS-dihydroperoxy 6-trans, 8-cis, 11-cis, 13-trans-eicosatetraenoic acid. Independent evidence for the absolute configurations was obtained by capillary gas-liquid chromatography of diastereomeric R-(-)-2-butyl esters of the acetylated 2-hydroxy carboxylic acids produced by oxidative ozonolysis of the acetylated dihydroxy fatty acids. It is concluded that soybean lipoxygenase-1 produces hydroperoxides with predominantly the S-configuration irrespective of the position in the fatty acid which is oxygenated.
Asunto(s)
Ácidos Araquidónicos , Lipooxigenasa/metabolismo , Plantas/enzimología , Araquidonato Lipooxigenasas , Ácidos Araquidónicos/síntesis química , Cinética , Espectroscopía de Resonancia Magnética , Espectrometría de Masas , Métodos , Glycine max , Espectrofotometría Ultravioleta , Especificidad por SustratoRESUMEN
The elucidation of the structures of two carbohydrate units, N-glycosidically linked to an asparagine residue of bovine lactotransferrin, is described. These carbohydrate structures are of the oligomannoside type and contain eight or nine mannose residues, respectively. The potency of 500 MHz 1H-NMR spectroscopy in primary structure determination of two closely related carbohydrate chains present in a mixture is demonstrated. This implies that 500 MHz 1H-NMR spectroscopy can disclose microheterogeneity which is almost untraceable using other approaches.