RESUMO
A fatal case of malignant atrophic papulosis (Degos disease) with optic nerve and spinal cord involvement is described. Magnetic resonance imaging (MRI) of the optic nerve showed abnormal signal enhancement on fat suppressed T1 weighted images after intravenous meglumine gadopentetate infusion. On T2 weighted sagittal images, a sawtooth pattern was observed over seven vertebral segments of the spinal cord. On necropsy, a severe loss of myelinated nerve fibres in the left optic nerve was seen, with thrombotic obstruction of the central retinal artery. Spongy degeneration was observed in all levels of the spinal cord, with patchy and motheaten patterns caused by thromboses and endothelial proliferation in subarachnoid vessels. Findings on MRI were consistent with findings on pathological examination.
Assuntos
Doenças do Nervo Óptico/diagnóstico , Pele/patologia , Doenças da Medula Espinal/diagnóstico , Atrofia , Isquemia Encefálica/diagnóstico , Isquemia Encefálica/patologia , Progressão da Doença , Evolução Fatal , Feminino , Humanos , Aumento da Imagem , Embolia Intracraniana/diagnóstico , Embolia Intracraniana/patologia , Imageamento por Ressonância Magnética , Pessoa de Meia-Idade , Degeneração Neural/diagnóstico , Degeneração Neural/patologia , Nervo Óptico/patologia , Doenças do Nervo Óptico/patologia , Dermatopatias Papuloescamosas/diagnóstico , Dermatopatias Papuloescamosas/patologia , Medula Espinal/patologia , Doenças da Medula Espinal/patologiaRESUMO
Positional cloning approaches revealed that Tangier disease (TD), a genetic high density lipoprotein deficiency, is associated with mutations in the ATP-binding cassette transporter-1 (ABCA1) gene. However, the biological function of ABCA1 is still not fully investigated. Recently, we have reported that the cells from the patients with TD had abnormal actin cytoskeletons in association with decreased expression of Cdc42, a member of RhoGTPases family. In the present study, we have found that actin cytoskeletons were altered in HEK293 cells transfected with human ABCA1 (hABCA1) cDNA. Cells expressing hABCA1 were divided into the following two groups by the distinct morphology with altered actin cytoskeletons: one had increased formation of filopodia (designated as Type I) and the other had long protrusions (designated as Type II). Type I cells had morphology similar to that of cells transfected with dominant active form of Cdc42 (Cdc42-DA, V12Cdc42Hs-DA). Type II cells had morphology similar to that of cells transfected with neural Wiskott-Aldrich Syndrome Protein (N-WASP),one of the established downstream effector molecules of Cdc42. We have obtained the data showing a possible pathway of ABCA1/Cdc42/N-WASP by the following experiments. Introduction of mutant of Cdc42 (dominant negative form of Cdc42, N17Cdc42Hs-DN) and N-WASP (N-WASP lacking verprolin homology domain, N-WASPDeltaVPH), both of which are supposed to have potential to inhibit rearrangement of actin cytoskeletons, significantly inhibited the morphological changes induced by expression of hABCA1. Immunoprecipitation study with FLAG-tagged ABCA1 (hABCA1-FLAG) revealed that Cdc42 was coimmunoprecipitated with hABCA1-FLAG. In addition, we have demonstrated possible intracellular colocalization of these two molecules in the overexpressing cells by the confocal laser microscopy. These results may suggest that hABCA1 regulates actin organization through the possible interaction with Cdc42Hs.
Assuntos
Transportadores de Cassetes de Ligação de ATP/metabolismo , Actinas/metabolismo , Citoesqueleto/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Proteína cdc42 de Ligação ao GTP/metabolismo , Transportador 1 de Cassete de Ligação de ATP , Animais , Western Blotting , Células COS , Linhagem Celular , Membrana Celular/metabolismo , DNA Complementar/metabolismo , Genes Dominantes , Humanos , Microscopia Confocal , Microscopia de Fluorescência , Plasmídeos/metabolismo , Testes de Precipitina , Ligação Proteica , Transfecção , Proteína Neuronal da Síndrome de Wiskott-Aldrich , Proteína cdc42 de Ligação ao GTP/genéticaRESUMO
Dipeptidyl peptidase III (DPP III) (EC 3.4.14.4), which has a HELLGH-E (residues 450-455, 508) motif as the zinc binding site, is classified as a zinc metallopeptidase. The zinc dissociation constants of the wild type, Leu(453)-deleted, and E508D mutant of DPP III at pH 7.4 were 4.5 (+/-0.7) x 10(-13), 5.8 (+/-0.7) x 10(-12), and 3.2 (+/-0.9) x 10(-10) M, respectively. The recoveries of the enzyme activities by the addition of various metal ions to apo-DPP III were also measured, and Co(2+), Ni(2+), and Cu(2+) ions completely recovered the enzyme activities as did Zn(2+). The dissociation constants of Co(2+), Ni(2+), and Cu(2+) ions for apo-DPP III at pH 7.4 were 8.2 (+/-0.9) x 10(-13), 2.7 (+/-0.3) x 10(-12), and 1.1 (+/-0.1) x 10(-14) M, respectively. The shape of the absorption spectrum of Co(2+)-DPP III was very similar to that of Co(2+)-carboxypeptidase A or Co(2+)-thermolysin, in which the Co(2+) is bound to two histidyl nitrogens, a water molecule, and a glutamate residue. The absorption spectrum of Cu(2+)-DPP III is also very similar to that of Cu(2+)-thermolysin. The EPR spectrum and the EPR parameters of Cu(2+)-DPP III were very similar to those of Cu(2+)-thermolysin but slightly different from those of Cu(2+)-carboxypeptidase A. The five lines of the superfine structure in the perpendicular region of the EPR spectrum in Cu(2+)-DPP III suggest that nitrogen atoms should coordinate to the cupric ion in Cu(2+)-DPP III. All of these data suggest that the donor set and the coordination geometry of the metal ions in DPP III, which has the HExxxH motif as the metal binding site, are very similar to those of the metal ions in thermolysin, which has the HExxH motif.
Assuntos
Dipeptidil Peptidases e Tripeptidil Peptidases/metabolismo , Fígado/enzimologia , Zinco/metabolismo , Animais , Espectroscopia de Ressonância de Spin Eletrônica , Cinética , Ratos , Proteínas Recombinantes/metabolismoRESUMO
Cholesterol efflux (CE) is the initial and important step of reverse cholesterol transport (RCT), a major protective system against atherosclerosis. However, most of the molecular mechanism for CE still remains to be clarified. In the present study, cDNA subtraction revealed that the expression of a member of the Rho GTPase family, Cdc42Hs, was markedly decreased in both passaged fibroblasts and macrophages (Mφ) from patients with Tangier disease (TD), a rare lipoprotein disorder with reduced CE. This small G protein is known to have many cell biological activities such as rearrangement of actin cytoskeleton and vesicular transport, however the association between this molecule and lipid transport has never been reported. We demonstrate that MDCK cells expressing the dominant negative form of Cdc42Hs had reduced CE, inversely ones expressing the dominant active form had increased CE. From these observations, we would like to raise a novel hypothesis that this type of small G protein may play a role in some steps of CE. To our knowledge, the present study is the first demonstration that the expression of this molecule is altered in cells from human disease.
Assuntos
Proteínas Tirosina Quinases/metabolismo , Doença de Tangier/enzimologia , Proteína cdc42 de Ligação ao GTP/metabolismo , Animais , Linhagem Celular , Células Cultivadas , Cães , Fibroblastos/enzimologia , Biblioteca Gênica , Humanos , Macrófagos/enzimologia , Masculino , Pessoa de Meia-Idade , Proteínas Tirosina Quinases/genética , Pele/enzimologia , Doença de Tangier/genética , Transfecção , Proteína cdc42 de Ligação ao GTP/genéticaAssuntos
Calcinose/complicações , Doença das Coronárias/complicações , Doença de Tangier/complicações , Medula Óssea/patologia , Calcinose/diagnóstico , Angiografia Coronária , Doença das Coronárias/diagnóstico , Humanos , Masculino , Pessoa de Meia-Idade , Doença de Tangier/patologia , Ultrassonografia de IntervençãoRESUMO
Human alpha-N-acetylgalactosaminidase (alpha-GalNAc; also known as alpha-galactosidase B) is the lysosomal exoglycohydrolase that cleaves alpha-N-acetylgalactosaminyl moieties in glycoconjugates. Mutagenesis studies indicated that the first five (N124, N177, N201, N359, and N385) of the six potential N-glycosylation sites were occupied. Site 3 occupancy was important for enzyme function and stability. Characterization of the N-linked oligosaccharide structures on the secreted enzyme overexpressed in Chinese hamster ovary cells revealed highly heterogeneous structures consisting of complex (approximately 53%), hybrid (approximately 12%), and high mannose-type (approximately 33%) oligosaccharides. The complex structures were mono-, bi-, 2,4-tri-, 2,6-tri-, and tetraantennary, among which the biantennary structures were most predominant (approximately 53%). Approximately 80% of the complex oligo-saccharides had a core-region fucose and 50% of the complex oligosaccharides were sialylated exclusively with alpha-2,3-linked sialic acid residues. The majority of hybrid type oligo-saccharides were GalGlcNAcMan(6)GlcNAc-Fuc(0-1)GlcNAc. Approximately 54% of the hybrid oligosaccharide were phosphorylated and one-third of these structures were further sialylated, the latter representing unique phosphorylated and sialylated structures. Of the high mannose oligosaccharides, Man(5-7)GlcNAc(2) were the predominant species (approximately 90%) and about 50% of the high mannose oligosaccharides were phosphorylated, exclusively as monoesters whose positions were determined. Comparison of the oligosaccharide structures of alpha-GalNAc and alpha-galactosidase A, an evolutionary-related and highly homologous exoglycosidase, indicated that alpha-GalNAc had more completed complex chains, presumably due to differences in enzyme structure/domains, rate of biosynthesis, and/or aggregation of the overexpressed recombinant enzymes.
Assuntos
Hexosaminidases/química , Oligossacarídeos/química , Animais , Células COS , Configuração de Carboidratos , Sequência de Carboidratos , Cromatografia de Afinidade , Hexosaminidases/genética , Hexosaminidases/metabolismo , Humanos , Dados de Sequência Molecular , Mutagênese Sítio-Dirigida , Oligossacarídeos/metabolismo , alfa-N-AcetilgalactosaminidaseRESUMO
Previously, we showed that lowering the growth temperature increased the level of eicosapentaenoic acid (EPA) in the phosphatidylcholine (PtdCho) of Caenorhabditis elegans. In this study, we investigated the molecular species composition of PtdCho of C. elegans, with an emphasis on EPA-containing species. C. elegans contained a substantial amount of 1,2-dipolyunsaturated fatty acid-containing PtdCho (1,2-diPUFA-PtdCho) species, such as arachidonic acid/EPA and EPA/EPA, which are unusual phospholipids in higher animals. The EPA/EPA-PtdCho content was significantly increased in C. elegans grown at a low temperature. To examine the possibility that the acyltransferase activity involved in the remodeling of phospholipids accounts for the production of 1,2-diPUFA-PtdCho, we investigated the substrate specificity of this enzyme in C. elegans and found that it did not exhibit a preference for saturated fatty acid for acylation to the sn-1 position of PtdCho. The efficacy of the esterification of EPA to the sn-1 position was almost equal to that of stearic acid. The lack of preference for a saturated fatty acid for acylation to the sn-1 position of PtdCho is thought to result in the existence of the unusual 1,2-diEPA-PtdCho in C. elegans.
Assuntos
Caenorhabditis elegans/metabolismo , Glicerilfosforilcolina/análogos & derivados , Acilação , Aciltransferases/metabolismo , Animais , Caenorhabditis elegans/crescimento & desenvolvimento , Coenzima A Ligases/metabolismo , Ácidos Graxos/análise , Glicerilfosforilcolina/biossíntese , Fosfatidilcolinas/biossíntese , Fosfatidilcolinas/química , Fosfatidiletanolaminas/biossíntese , Fosfatidiletanolaminas/química , Especificidade por Substrato , TemperaturaRESUMO
The scavenger receptor class B type I (SR-BI) and its human homologue CLA-1 (CD36 and LIMPII Analogous-1) have recently been identified to bind HDL and mediate the selective uptake of HDL lipids. Tissue distribution of both murine and human receptors is quite similar, in that they are expressed abundantly in liver and steroidogenic tissues. However, expression and function of the human SR-BI (hSR-BI), in the periphery of reverse cholesterol transport such as macrophages, are still unclear. In the present study, we have raised two different kinds of anti-hSR-BI polypeptide antibodies (Abs): one against the extracellular domain and the other against the intracellular domain. We have investigated the expression of hSR-BI mRNA and immunoreactive mass in freshly isolated cultured human monocyte-derived macrophages (hMphi) and in atherosclerotic lesions. Contrary to the earlier report, hSR-BI mRNA was expressed in cultured hMphi and markedly upregulated with differentiation, determined by Northern blot and reverse transcriptase-based polymerase chain reaction analyses. The mRNA expression pattern during differentiation of hMphi was very similar to those of SR class A and another member of SR class B, CD36. Protein expression was confirmed by Western blot analyses with the above Abs to show a major 83-kDa band. Modified lipoproteins such as oxidized LDL and acetylated LDL induced a 5-fold increase in mRNA and protein expression of hSR-BI. Confocal immunofluorescence microscopy demonstrated that hSR-BI immunoreactive mass was detectable as a heterogeneous, punctate staining pattern. Furthermore, immunohistochemical analysis showed that immunoreactive mass of hSR-BI was detected in foam cells in human aortic atherosclerotic lesions and that there was no significant difference of staining patterns between the two Abs. This study clearly demonstrates that hSR-BI is expressed in the lipid-laden macrophages in human atherosclerotic lesions, suggesting that it is very important to know its function and regulation in hMphi to understand the biological utility of this molecule.
Assuntos
Arteriosclerose/metabolismo , Macrófagos/metabolismo , Proteínas de Membrana , Monócitos/citologia , Receptores Imunológicos/metabolismo , Receptores de Lipoproteínas , Animais , Aorta/metabolismo , Aorta/patologia , Arteriosclerose/patologia , Antígenos CD36 , Células CHO , Diferenciação Celular/fisiologia , Linhagem Celular , Cricetinae , Imunofluorescência , Humanos , Imuno-Histoquímica , Técnicas Imunológicas , Lipoproteínas/farmacologia , Macrófagos/efeitos dos fármacos , Microscopia de Fluorescência , Isoformas de Proteínas/metabolismo , RNA Mensageiro/metabolismo , Receptores Imunológicos/genética , Receptores Depuradores , Valores de Referência , Receptores Depuradores Classe A , Receptores Depuradores Classe BRESUMO
Necrobiotic xanthogranuloma (NXG) is a rare histiocytic disease with generalized xanthomatosis. However, most cases with NXG are normolipidemic or hypolipidemic. The mechanism for the formation of xanthoma in NXG has not yet been clarified. We observed a case of NXG with severe hypocholesterolemia (total cholesterol: 1.69 mmol/l) and analyzed the function of monocytes in this case. Histological examinations by light microscopy revealed a large amount of lipid deposition in the patient's freshly isolated monocytes. The patient's monocytes showed a 3-fold increase in cholesteryl ester content and a 3-fold enhancement of acetyl low density lipoprotein (LDL) uptake compared with the control monocytes. However, no significant difference was noted in the expression of CD36 protein and the mRNA levels of scavenger receptor-class A (SR-A) between the monocytes of the patient and the control. The phagocytotic ability of the patient's monocytes was enhanced 1.5-fold compared with that of the control monocytes. These findings suggest that the activated monocytes may have degraded the modified LDL via a pathway other than CD36 or SR-A, and accumulated a great amount of lipids in vivo. In conclusion, the present study has demonstrated a possible pathogenesis of NXG that the activation of monocytes in vivo may contribute to the intracellular accumulation of lipoprotein-derived lipids leading to non-inherited xanthomatosis and the marked hypocholesterolemia.
Assuntos
Colesterol/deficiência , Granuloma/etiologia , Lipoproteínas LDL/metabolismo , Proteínas de Membrana , Monócitos/fisiologia , Receptores de Lipoproteínas , Xantomatose/etiologia , Idoso , Animais , Antígenos CD36/genética , Antígenos CD36/metabolismo , Células Cultivadas , Colesterol/metabolismo , Ésteres do Colesterol/metabolismo , Primers do DNA/química , Fibroblastos/metabolismo , Fibroblastos/patologia , Fator Estimulador de Colônias de Granulócitos e Macrófagos/metabolismo , Granuloma/sangue , Granuloma/patologia , Humanos , Linfócitos/fisiologia , Masculino , Camundongos , Fagocitose , RNA Mensageiro/biossíntese , Receptores Imunológicos/genética , Receptores Imunológicos/metabolismo , Receptores Depuradores , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Receptores Depuradores Classe A , Receptores Depuradores Classe B , Xantomatose/sangue , Xantomatose/patologiaRESUMO
We investigated the relationship between uric acid (UA) metabolism and fat distribution in 36 obese men with a mean +/- SD age of 38 +/- 16 years and mean body-mass index (BMI) of 34 +/- 4 kg/m2. Subjects were divided into two groups: subcutaneous fat obesity (SFO) and visceral fat obesity (VFO), according to their abdominal fat distribution based on the results of computed tomography (CT). SFO was defined as having a ratio of visceral fat area (VFA) to subcutaneous fat area (V/S) of less than 0.4, and VFO was defined as having a V/S ratio > or = 0.4. The levels of serum total cholesterol (T-Chol), triglyceride (TG), and fasting plasma glucose (FPG), and the diastolic blood pressure (dBP) were significantly higher in the VFO group than in the SFO group. Serum UA levels were much higher in both the SFO and VFO groups than in the non-obese control group (492 +/- 107 and 474 +/- 90 v 309 +/- 48 micromol/L, respectively). The 24-hour urinary urate excretion (u-UA24h) and the UA clearance (Cua) to creatinine clearance (Ccr) ratio were significantly higher in the VFO group than in the SFO group (3.75 +/- 1.43 v 2.69 +/- 1.12 mmol/d, P < .05; and 5.9% +/- 2.0% v 3.6% +/- 1.7%, P < .001, respectively). The frequency of hyperuricemia was markedly higher in both the SFO and VFO groups compared with the control group (71% and 73% v 0%, respectively). Although the high serum UA level seemed to be related to low u-UA24h in 80% of SFO subjects with hyperuricemia, this was the case in only 10% of VFO subjects. While 44% of VFO subjects with hyperuricemia were designated as an overproduction type. These results suggest that the mechanism of hyperuricemia in obesity may be affected by the difference in body fat distribution and that the assessment of body fat distribution and types of hyperuricemia is crucial for the treatment of obese patients with hyperuricemia.
Assuntos
Tecido Adiposo/metabolismo , Obesidade/metabolismo , Pele , Ácido Úrico/metabolismo , Vísceras , Tecido Adiposo/patologia , Adolescente , Adulto , Idoso , Constituição Corporal , Índice de Massa Corporal , Humanos , Masculino , Pessoa de Meia-IdadeRESUMO
Human alpha-galactosidase A (alpha-Gal A) is the lysosomal glycohydrolase that cleaves the terminal alpha-galactosyl moieties of various glycoconjugates. Overexpression of the enzyme in Chinese hamster ovary (CHO) cells results in high intracellular enzyme accumulation and the selective secretion of active enzyme. Structural analysis of the N -linked oligosaccharides of the intracellular and secreted glycoforms revealed that the secreted enzyme's oligosaccharides were remarkably heterogeneous, having high mannose (63%), complex (30%), and hybrid (5%) structures. The major high mannose oligosaccharides were Man5-7GlcNAc2 species. Approximately 40% of the high mannose and 30% of the hybrid oligosaccharides had phosphate monoester groups. The complex oligosaccharides were mono-, bi-, 2,4-tri-, 2,6-tri- and tetraantennary with or without core-region fucose, many of which had incomplete outer chains. Approximately 30% of the complex oligosaccharides were mono- or disialylated. Sialic acids were mostly N -acetylneuraminic acid and occurred exclusively in alpha2, 3-linkage. In contrast, the intracellular enzyme had only small amounts of complex chains (7.7%) and had predominantly high mannose oligosaccharides (92%), mostly Man5GlcNAc2 and smaller species, of which only 3% were phosphorylated. The complex oligosaccharides were fucosylated and had the same antennary structures as the secreted enzyme. Although most had mature outer chains, none were sialylated. Thus, the overexpression of human alpha-Gal A in CHO cells resulted in different oligosaccharide structures on the secreted and intracellular glycoforms, the highly heterogeneous secreted forms presumably due to the high level expression and impaired glycosylation in the trans- Golgi network, and the predominately Man5-7GlcNAc2 cellular glycoforms resulting from carbohydrate trimming in the lysosome.
Assuntos
Oligossacarídeos/química , alfa-Galactosidase/química , Animais , Células CHO , Sequência de Carboidratos , Cricetinae , Expressão Gênica , Glicoconjugados/metabolismo , Glicosilação , Humanos , Lisossomos/enzimologia , Dados de Sequência Molecular , Estrutura Molecular , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , alfa-Galactosidase/genética , alfa-Galactosidase/metabolismoRESUMO
We demonstrated a continuous intracoronary infusion of acetylcholine-induced marked decrease of coronary blood flow estimated by intracoronary Doppler flow wire without significant epicardial coronary narrowing. This case can be called a patient with microvascular vasospastic angina.
Assuntos
Acetilcolina/efeitos adversos , Angina Pectoris/induzido quimicamente , Acetilcolina/administração & dosagem , Angina Pectoris/diagnóstico por imagem , Angina Pectoris/fisiopatologia , Angiografia Coronária , Vasos Coronários/diagnóstico por imagem , Ecocardiografia Doppler , Eletrocardiografia , Feminino , Humanos , Infusões Intra-Arteriais , Pessoa de Meia-IdadeRESUMO
Lysosomal storage diseases associated with early-onset pathological changes may require prenatal therapy to avert the profound effects of the metabolic error on organs, especially the central nervous system. The present investigation determined the extent of expression of beta-mannosidase deficiency in the caprine fetus at 62 days of gestation, near the end of the period of immunotolerance when donor cells can engraft in various organs without immune rejection and supply missing enzyme. Three pairs of obligate carrier goats from the beta-mannosidosis colony were mated. Out of six fetuses delivered at 62 days of gestation, one (V385) was identified by measurement of beta-mannosidase activity as the only fetus affected with beta-mannosidosis. Thin-layer chromatography and quantitation of oligosaccharides revealed the presence of tri- and disaccharides, typical of beta-mannosidosis, only in V385. Morphological analysis revealed cytoplasmic vacuolation typical of beta-mannosidosis in V385; in thyroid, spinal cord, and kidney, the pattern of vacuolation was similar to, but less severe than, that observed previously in newborn affected goats. On the basis of these results, it will be possible to determine the effects of prenatal cell transplantation therapeutic strategies performed during the period of immunotolerance by monitoring phenotypic characteristics after treatment.
Assuntos
Doenças Fetais/veterinária , Doenças das Cabras/metabolismo , Cabras , alfa-Manosidose/veterinária , Animais , Encéfalo/enzimologia , Progressão da Doença , Feminino , Doenças Fetais/metabolismo , Doenças Fetais/patologia , Doenças Fetais/terapia , Idade Gestacional , Doenças das Cabras/patologia , Tolerância Imunológica , Rim/enzimologia , Masculino , Manosidases/metabolismo , Oligossacarídeos/metabolismo , alfa-Manosidose/metabolismo , alfa-Manosidose/patologia , alfa-Manosidose/terapia , beta-ManosidaseRESUMO
A study was conducted on calcium chloride treatments of canned okra acidified by adding either acetic, citric, lactic, malic or tartaric acids or by lactic fermentation. The quality of the processed okra was determined by physical, chemical, microbiological and sensory analyses after a two month storage period at room temperature. The results indicated the possibility of processing high quality canned okra by small canneries, with low cost equipment and low energy requirements. The acidification procedures ensure minimal risk of botulism.
Assuntos
Cloreto de Cálcio/administração & dosagem , Conservação de Alimentos/normas , Malvaceae/química , Ácido Acético/administração & dosagem , Ácido Acético/efeitos adversos , Botulismo/prevenção & controle , Cloreto de Cálcio/efeitos adversos , Ácido Cítrico/administração & dosagem , Ácido Cítrico/efeitos adversos , Manipulação de Alimentos/normas , Concentração de Íons de Hidrogênio , Ácido Láctico/administração & dosagem , Ácido Láctico/efeitos adversos , TemperaturaRESUMO
A study was conducted on calcium chloride treatment of canned okra acidified by adding either acetic, lactic, malic or tartaric acids or by lactic fermentation. The quality of the processed okra was determined by physical, chemical, microbiological and sensory analyses after a two month storage period at room temperature. The results indicated the possibility of processing high quality okra by small canneries, with low cost equipment and low energy requirements. The acidification procedures ensure minimal risk of botulism
Assuntos
Humanos , Botulismo , Cálcio/administração & dosagem , Cálcio/classificação , Cálcio/uso terapêuticoRESUMO
Ascorbate oxidase from the fungus Acremonium sp. HI-25 is a copper-containing glycoprotein that catalyzes the oxidation of ascorbic acid to dehydroascorbic acid. Monosaccharide composition analysis showed that the enzyme contains exclusively N-linked oligosaccharide chains. Following liberation by hydrazinolysis/re-N-acetylation, and fractionation by HPLC on anion exchange. Amide-80 and/or octadecyl silica columns after derivatization with p-aminobenzoic ethyl ester, the structures of the twelve major neutral oligosaccharides were identified by FAB-MS, 400 MHz 1H-NMR, methylation analysis, mild acid hydrolysis, and/or sequential exoglycosidase digestions. Acremonium sp. ascorbate oxidase was found to consist of high-mannose type oligosaccharides (76.3%) having 4 to 9 mannose residues and a series of novel D-galactofuranose-containing high-mannose type oligosaccharides (18.6%) with the following structure.
Assuntos
Acremonium/enzimologia , Ascorbato Oxidase/metabolismo , Manose/metabolismo , Oligossacarídeos/metabolismo , Acremonium/metabolismo , Ascorbato Oxidase/química , Sequência de Carboidratos , Cromatografia Líquida de Alta Pressão , Cromatografia por Troca Iônica , Galactose/química , Galactose/metabolismo , Espectroscopia de Ressonância Magnética , Manose/química , Dados de Sequência Molecular , Oligossacarídeos/química , Espectrometria de Massas de Bombardeamento Rápido de ÁtomosRESUMO
The phospholipid and the fatty chain compositions of diacyl, alkylacyl and alkenylacyl glycerophospholipids of the free-living nematode, Caenorhabditis elegans, were investigated. The phospholipids were comprised of 54.5% ethanolamine glycerophospholipid (EGP), 32.3% choline glycerophospholipid (CGP), 8.1% sphingomyelin and 5.1% others. The most abundant fatty acid in CGP was eicosapentaenoic acid (20:5n-3). The fatty acids in CGP were more unsaturated than those in EGP. Alkenylacyl and alkylacyl subclasses accounted for 1.0 and 2.6%, respectively, of CGP and 14.0 and 19.6%, respectively, of EGP. At least 80% of the alkenyl and alkyl groups were 18:0 chains and the remaining were odd numbered chains. The potential presence of platelet-activating factor (PAF) was examined by bioassay, but PAF-like activity was not detected in the extracts of this nematode.
Assuntos
Caenorhabditis elegans/química , Fosfolipídeos/análise , Animais , Bioensaio , Escherichia coli , Ácidos Graxos/análise , Fosfatidilcolinas/análise , Fosfatidiletanolaminas/análise , Fator de Ativação de Plaquetas/análiseRESUMO
Structures of the Asn linked oligosaccharides of quail egg-yolk immunoglobulin (IgY) were determined in this study. Asn linked oligosaccharides were cleaved from IgY by hydrazinolysis and labelled with p-aminobenzoic acid ethyl ester (ABEE) after N-acetylation. The ABEE labelled oligosaccharides were then fractionated by a combination of Concanavalin A-agarose column chromatography and anion exchange, normal phase and reversed phase HPLC before their structures were determined by sequential exoglycosidase digestion, methylation analysis, HPLC, and 500 MHz 1H-NMR spectroscopy. Quail IgY contained only neutral oligosaccharides of the following categories: the glucosylated oligomannose type (0.6% Glc alpha 1-3Glc alpha 1-3Man9GlcNAc2; 35.6%, Glc alpha 1-3Man7-9GlcNAc2). oligomannose type (15.0%, with the structure Man5-9GlcNAc2) and biantennary complex type with core structures of -Man alpha 1-3(-Man alpha 1-6)Man beta 1-4GlcNAc beta 1-4GlcNAc (9.9%), -Man alpha 1-3 (GlcNAc beta 1-4)(-Man alpha 1-6)Man beta 1-4GlcNAc beta 1-4GlcNAc (25.1%) and -Man alpha 1-3(GlcNAc beta 1-4)(-Man alpha 1-6)Man beta 1-4GlcNAc beta 1-4(Fuc alpha 1-6)GlcNAc (11.4%). Although never found in mammalian proteins, glucosylated oligosaccharides (Glc1Man7-9GlcNAc2) have been located previously in hen IgY.