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1.
Appl Microbiol Biotechnol ; 65(5): 583-92, 2004 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-15248038

RESUMO

Arthrobacter globiformis T6 is unique in that it produces an enzyme yielding only isomaltose from dextran. In the present study, the organism was re-identified and its classification as a new species of the genus Arthrobacter, A. dextranlyticum, was proposed. The high G+C gene (66.8 mol%) for the isomalto-dextranase was sequenced. The deduced amino acid sequence, with a calculated molecular mass of 65,993 Da (603 amino acids), was confirmed by nanoscale capillary liquid chromatography coupled to tandem mass spectrometry, which covered 71.1% of the amino acid residues of the entire sequence. The enzyme was grouped into glycoside hydrolase family 27, and the C-terminal domain has homology to carbohydrate-binding module family 6. Hyper-exoproduction of the recombinant enzyme was achieved at a level corresponding to approximately 4.6 g l(-1) of culture broth when proteases-deficient Bacillus subtilis cells were used as the host. The purified enzyme (65.5 kDa) had an optimal pH and temperature for activity of 3.5 and 60 degrees C, respectively. It was crystallized using the sitting-drop vapor-diffusion method at 293 K.


Assuntos
Arthrobacter/enzimologia , Bacillus subtilis/genética , Glicosídeo Hidrolases/genética , Glicosídeo Hidrolases/metabolismo , Peptídeo Hidrolases/genética , Sequência de Aminoácidos , Arthrobacter/classificação , Arthrobacter/genética , Bacillus subtilis/enzimologia , Bacillus subtilis/metabolismo , Composição de Bases , Sequência de Bases , Clonagem Molecular , Cristalização , DNA Bacteriano/química , DNA Bacteriano/isolamento & purificação , DNA Ribossômico/química , DNA Ribossômico/isolamento & purificação , Estabilidade Enzimática , Deleção de Genes , Expressão Gênica , Genes Bacterianos , Genes de RNAr/genética , Glicosídeo Hidrolases/química , Concentração de Íons de Hidrogênio , Dados de Sequência Molecular , Peso Molecular , Filogenia , Estrutura Terciária de Proteína , RNA Bacteriano/genética , RNA Ribossômico 16S/genética , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/isolamento & purificação , Proteínas Recombinantes/metabolismo , Análise de Sequência de DNA , Temperatura
2.
Appl Microbiol Biotechnol ; 64(4): 505-14, 2004 May.
Artigo em Inglês | MEDLINE | ID: mdl-15088129

RESUMO

An agar-degrading bacterium, strain JAMB-A7, was isolated from the sediment in Sagami Bay, Japan, at a depth of 1,174 m and identified as a novel species of the genus Microbulbifer. The gene for a novel beta-agarase from the isolate was cloned and sequenced. It encodes a protein of 441 amino acids with a calculated molecular mass of 48,989 Da. The deduced amino acid sequence showed similarity to those of known beta-agarases in glycoside hydrolase family 16, with only 34-55% identity. A sequence similar to a carbohydrate-binding module was found in the C-terminal region of the enzyme. The recombinant agarase was hyper-produced extracellularly using Bacillus subtilis as the host, and the enzyme purified to homogeneity had a specific activity of 398 U (mg protein)(-1) at pH 7.0 and 50 degrees C. It was thermostable, with a half-life of 502 min at 50 degrees C. The optimal pH and temperature for activity were around 7 and 50 degrees C, respectively. The pattern of agarose hydrolysis showed that the enzyme was an endo-type beta-agarase, and the final main product was neoagarotetraose. The activity was not inhibited by NaCl, EDTA, and various surfactants at high concentrations. In particular, sodium dodecyl sulfate had no inhibitory effect up to 2%.


Assuntos
Alteromonadaceae/enzimologia , Glicosídeo Hidrolases/genética , Glicosídeo Hidrolases/metabolismo , Alteromonadaceae/classificação , Alteromonadaceae/isolamento & purificação , Sequência de Aminoácidos , Bacillus subtilis/genética , Bacillus subtilis/metabolismo , Sequência de Bases , Clonagem Molecular , Códon de Terminação/genética , DNA Bacteriano/química , DNA Bacteriano/isolamento & purificação , DNA Ribossômico/química , DNA Ribossômico/isolamento & purificação , Estabilidade Enzimática , Galactosídeos/análise , Galactosídeos/metabolismo , Genes Bacterianos , Genes de RNAr/genética , Sedimentos Geológicos/microbiologia , Glicosídeo Hidrolases/química , Glicosídeo Hidrolases/isolamento & purificação , Concentração de Íons de Hidrogênio , Dados de Sequência Molecular , Oligossacarídeos/análise , Oligossacarídeos/metabolismo , Fases de Leitura Aberta , Regiões Promotoras Genéticas , Sinais Direcionadores de Proteínas/genética , RNA Ribossômico 16S/genética , Proteínas Recombinantes/isolamento & purificação , Proteínas Recombinantes/metabolismo , Sequências Repetitivas de Ácido Nucleico , Análise de Sequência de DNA , Homologia de Sequência de Aminoácidos , Temperatura , Microbiologia da Água
3.
Microb Ecol ; 47(2): 186-96, 2004 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-14749907

RESUMO

Massive chimney structures, which are characteristic of many hydrothermally active zones, harbor diverse microbial communities containing both thermophilic and hyperthermophilic microbes. However, vent chimneys ultimately become hydrothermally inactive, and the changes that occur in the microbial communities upon becoming inactive have not been documented. We thus collected inactive chimneys from two geologically and geographically distinct hydrothermal fields, Iheya North in the western Pacific Ocean and the Kairei field in the Indian Ocean. The chimneys displayed easily distinguishable strata, which were analyzed with regard to both mineralogical and microbiological properties. X-ray diffraction pattern and energy-dispersive spectroscopic analyses revealed that the main mineral components of the chimney substructures from Iheya North and the Kairei field were barite (BaSO4) and chalcopyrite (CuFeS2), respectively. Microbial cell densities in the substructures determined by DAPI counting ranged from 1.7 x 10(7) cells g(-1) to 3.0 x 10(8) cells g(-1). The proportions of archaeal rDNA in the whole microbial rDNA assemblages in all substructures were, at most, a few percent as determined by quantitative fluorogenic PCR. The microbial rDNA clone analysis and whole-cell fluorescence in situ hybridization revealed a community that was decidedly different from any communities previously reported in active chimneys. Curiously, both samples revealed the abundant presence of a group of Bacteria related to a magnetosome-bearing bacterium, " Magnetobacterium bavaricum" of the Nitrospirae division. These results suggest that inactive chimneys provide a distinct microbial habitat.


Assuntos
Archaea/genética , Bactérias/genética , Biodiversidade , Ecossistema , Sedimentos Geológicos/microbiologia , Filogenia , Sulfato de Bário , Sequência de Bases , Contagem de Colônia Microbiana , Cobre , Primers do DNA , DNA Ribossômico/genética , Sedimentos Geológicos/análise , Hibridização in Situ Fluorescente , Indóis , Dados de Sequência Molecular , Oceanos e Mares , Reação em Cadeia da Polimerase , Análise de Sequência de DNA , Análise Espectral , Difração de Raios X
4.
Pediatr Surg Int ; 17(5-6): 386-9, 2001 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-11527172

RESUMO

Inchin-ko-to (ICKT) prevents Fas-mediated liver injury. This study evaluates the effect of ICKT on conventional markers of liver function (LF) and liver fibrosis in 18 postoperative biliary atresia (BA) patients aged 3 to 23 years with elevated glutamic oxaloacetic transaminase (GOT), glutamic pyruvic transaminase (GPT), gamma-glutamyl transpeptidase (gammaGTP) but normal serum total bilirubin (T-Bil) levels. ICKT (0.15 g/kg per day) was administered orally for 1 year. Serum GOT, GPT, gammaGTP, total bile acids (TBA), and T-Bil as markers of LF and hyaluronic acid (HA), prolyl hydroxylase (PH), procollagen III peptide (PIIIP), and type IV collagen as markers of liver fibrosis were measured before and after treatment in each patient and compared statistically. All patients tolerated ICKT well, and there were no side effects. The percentage of subjects who improved after ICKT was 45% for serum GOT, 72% for GPT, 72% for gammaGTP, 72% for TBA, 67% for HA, 40% for PH, 50% for PIIIP, and 23% for type IV collagen. Changes in the mean values of all serum markers were statistically significant (P < 0.01). It is concluded that long-term administration of ICKT in postoperative BA patients improves liver status as assessed by markers of LF and fibrosis.


Assuntos
Atresia Biliar/cirurgia , Medicamentos de Ervas Chinesas/uso terapêutico , Cirrose Hepática/prevenção & controle , Inibidores de Fosfodiesterase/uso terapêutico , Fitoterapia , Portoenterostomia Hepática , Complicações Pós-Operatórias/prevenção & controle , Adolescente , Adulto , Atresia Biliar/mortalidade , Criança , Pré-Escolar , Humanos , Japão/epidemiologia , Testes de Função Hepática , Cuidados Pós-Operatórios , Taxa de Sobrevida
5.
Int J Syst Evol Microbiol ; 51(Pt 4): 1425-1435, 2001 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-11491342

RESUMO

A novel extreme thermophile was isolated from a water sample derived from a deep subsurface geothermal water pool at a depth of 1500 m in the Hacchoubaru geothermal plant in Oita Prefecture, Japan. The cells were found to be straight rods, each being motile by means of a polar flagellum. Growth was observed at temperatures between 60 and 85 degrees C (optimum 78 degrees C; 120 min doubling time) and between pH 5.5 and pH 9.0 (optimum 7.5). The isolate was a strictly aerobic heterotroph capable of utilizing a number of substrates such as yeast extract, peptone, tryptone, various carbohydrates, sugars, amino acids and organic acids. Elemental sulfur, thiosulfate, sulfide or cysteine-hydrochloride was required as an electron donor for growth. Hydrogen gas did not support growth. The G+C content of the genomic DNA was 44.7 mol%. Phylogenetic analysis based on 16S rDNA sequences and DNA-DNA hybridization analysis indicated that the isolate was closely related to members of the hydrogen-oxidizing, autotrophic and thermophilic genera Hydrogenobacter and Calderobacterium. However this isolate was differentiated from the previously described species of these genera on the basis of the physiological and molecular properties of the new isolate. The name Hydrogenobacter subterraneus sp. nov. is proposed; the type strain is HGP1T (= JCM 10560T = IFO 16485T).


Assuntos
Bactérias/classificação , Bactérias/isolamento & purificação , Microbiologia da Água , Bactérias/genética , Bactérias/crescimento & desenvolvimento , Bactérias/metabolismo , Composição de Bases , DNA Bacteriano/química , DNA Bacteriano/genética , DNA Ribossômico/genética , Ácidos Graxos/análise , Temperatura Alta , Hidrogênio/metabolismo , Japão , Microscopia Eletrônica , Dados de Sequência Molecular , Hibridização de Ácido Nucleico , Filogenia , Especificidade da Espécie , Terminologia como Assunto
6.
J Pediatr Surg ; 36(8): 1297-301, 2001 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-11479880

RESUMO

BACKGROUND/PURPOSE: Biliary atresia (BA) is associated with progressive liver fibrosis, which may be mediated by immunologic abnormalities involving adhesion molecules. This study investigates the relationship between serum intercellular adhesion molecule-1 (sICAM-1), serum vascular cell adhesion molecule-1 (sVCAM-1), and the clinical and histologic severity of BA. METHODS: Serum ICAM-1 and VCAM-1 levels were measured by enzyme-linked immunosorbent assay in 35 patients with BA and 20 healthy controls. Standard liver function tests (LFTs), and frozen section liver biopsy specimens were used to determine liver status. On the basis of LFT results, the BA patients were classified into group I (n = 10; normal LFTs), group II (n = 15; elevated LFTs, anicteric), and group III (n = 10; elevated LFTs, icteric). Eight subjects in group II, and all subjects in group III had portal hypertension (PH). RESULTS: sICAM-1 levels were significantly elevated in group III (1760.0 +/- 717.5 ng/mL) compared with group II (555.1 +/- 199.4 ng/mL), group I (272.1 +/- 59.9 ng/mL) and controls (256.3 +/- 71.6 ng/mL). Although sVCAM-1 levels were significantly elevated in group III (1932.9 +/- 282.6 ng/mL) compared with group II (1054.3 +/- 297.0 ng/mL), group I (605.4 +/- 112.4 ng/mL), and controls (616.0 +/- 112.0 ng/mL; P <.001), there was no statistically significant difference between groups I, II, or controls. sVCAM-1 levels were elevated significantly in BA subjects in group II with PH (1253.0 +/- 245.1 ng/mL) compared with those who did not have PH (827.3 +/- 151.7 ng/mL; P <.01). PH did not affect sICAM-1 levels. There was strong expression of ICAM-1 and VCAM-1 in proliferating bile ductules, endothelial cells, and liver cells in group III compared with group II and controls. CONCLUSIONS: In BA, sICAM-1 and sVCAM-1 levels could be useful as markers of end-stage liver disease, with sVCAM-1 being more specific for PH. Induction of ICAM-1 and VCAM-1 may be an important factor in the development of cirrhosis.


Assuntos
Atresia Biliar/sangue , Molécula 1 de Adesão Intercelular/sangue , Cirrose Hepática/sangue , Molécula 1 de Adesão de Célula Vascular/sangue , Adolescente , Atresia Biliar/complicações , Atresia Biliar/patologia , Atresia Biliar/cirurgia , Biomarcadores/análise , Biópsia por Agulha , Criança , Ensaio de Imunoadsorção Enzimática , Feminino , Humanos , Imuno-Histoquímica , Molécula 1 de Adesão Intercelular/análise , Fígado/química , Fígado/patologia , Cirrose Hepática/complicações , Cirrose Hepática/patologia , Testes de Função Hepática , Masculino , Período Pós-Operatório , Prognóstico , Valores de Referência , Sensibilidade e Especificidade , Índice de Gravidade de Doença , Molécula 1 de Adesão de Célula Vascular/análise
7.
Eur J Pediatr Surg ; 11(3): 163-6, 2001 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-11475111

RESUMO

Urinary Tyrosine Inhibitor (UTI) is produced in the liver and excreted in urine hepatic inflammation, infection or malignancy. We assess the possible implications of UTI in biliary atresia (BA). Liver function was used to divide 34 postoperative BA patients into 2 groups: Group 1 (n=25), anicteric (total bilirubin [T-Bil] <2.0 mg/dl); and Group 2 (n = 9), icteric (total bilirubin >2.0 mg/dl) with abnormal liver function test results, and repeated episodes of cholangitis. 26 age-matched subjects with no history of liver disease acted as controls


Assuntos
Atresia Biliar/complicações , Atresia Biliar/cirurgia , Glicoproteínas/metabolismo , Cirrose Hepática/etiologia , Cirrose Hepática/patologia , Adolescente , Atresia Biliar/diagnóstico , Biomarcadores/urina , Estudos de Casos e Controles , Criança , Feminino , Glicoproteínas/análise , Humanos , Imuno-Histoquímica , Testes de Função Hepática , Masculino , Monitorização Fisiológica/métodos , Complicações Pós-Operatórias , Probabilidade , Prognóstico , Radioimunoensaio , Valores de Referência , Sensibilidade e Especificidade
8.
Appl Environ Microbiol ; 67(8): 3618-29, 2001 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-11472939

RESUMO

Archaeal community structures in microhabitats in a deep-sea hydrothermal vent chimney structure were evaluated through the combined use of culture-independent molecular analyses and enrichment culture methods. A black smoker chimney was obtained from the PACMANUS site in the Manus Basin near Papua New Guinea, and subsamples were obtained from vertical and horizontal sections. The elemental composition of the chimney was analyzed in different subsamples by scanning electron microscopy and energy-dispersive X-ray spectroscopy, indicating that zinc and sulfur were major components while an increased amount of elemental oxygen in exterior materials represented the presence of oxidized materials on the outer surface of the chimney. Terminal restriction fragment length polymorphism analysis revealed that a shift in archaeal ribotype structure occurred in the chimney structure. Through sequencing of ribosomal DNA (rDNA) clones from archaeal rDNA clone libraries, it was demonstrated that the archaeal communities in the chimney structure consisted for the most part of hyperthermophilic members and extreme halophiles and that the distribution of such extremophiles in different microhabitats of the chimney varied. The results of the culture-dependent analysis supported in part the view that changes in archaeal community structures in these microhabitats are associated with the geochemical and physical dynamics in the black smoker chimney.


Assuntos
Archaea/classificação , Archaea/crescimento & desenvolvimento , Ecossistema , Água do Mar/microbiologia , Archaea/genética , Contagem de Colônia Microbiana , Meios de Cultura , DNA Ribossômico/análise , Temperatura Alta , Dados de Sequência Molecular , Filogenia , Reação em Cadeia da Polimerase , Polimorfismo de Fragmento de Restrição , RNA Ribossômico/genética , Ribotipagem , Água do Mar/química , Análise de Sequência de DNA , Análise Espectral
9.
Jpn J Pharmacol ; 85(4): 453-6, 2001 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-11388650

RESUMO

To investigate the mechanism for the amelioration by olopatadine hydrochloride (olopatadine) of allergic rhinitis, we determined its effects on the increase of chemical mediator concentrations in nasal lavage fluid following the intranasal antigen challenge in guinea pigs actively sensitized with DNP-Ascaris. The concentrations of histamine and peptide-leukotrienes increased 10 min after the challenge. Olopatadine at 10 mg/kg (p.o.) significantly prevented the increase of histamine and tended to inhibit the increase of peptide-leukotrienes. The inhibition by olopatadine of the nasal symptoms seems to involve the inhibitory effect on the releases of histamine and, possibly, p-LTs into the nasal cavity.


Assuntos
Anticorpos Anti-Helmínticos/metabolismo , Reações Antígeno-Anticorpo/efeitos dos fármacos , Antígenos de Helmintos/administração & dosagem , Antígenos de Helmintos/imunologia , Dibenzoxepinas/farmacologia , Antagonistas dos Receptores Histamínicos H1/farmacologia , Leucotrienos/metabolismo , Líquido da Lavagem Nasal/imunologia , Animais , Reações Antígeno-Anticorpo/imunologia , Antígenos de Helmintos/metabolismo , Cobaias , Imunidade Ativa , Injeções Intraperitoneais , Masculino , Testes de Provocação Nasal/métodos , Cloridrato de Olopatadina , Tromboxano B2/metabolismo
10.
Biosci Biotechnol Biochem ; 65(3): 690-3, 2001 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-11330692

RESUMO

We have isolated the genes for quinol oxidase from a deep-sea piezophilic bacterium, Shewanella violacea. Analysis of the deduced amino acid sequences of the cyo subunits showed that this oxidase has high similarity to Escherichia coli bo-type quinol oxidase. Northern blot analysis showed that these genes are expressed at a high level when the bacterium is grown at elevated pressure. Upstream in the cyo-operon, a sigma54-binding motif and an octamer sequence unit were found, suggesting that these elements may play a role in regulation of expression of the cyo-operon in response to changes in pressure.


Assuntos
Óperon , Oxirredutases/genética , Shewanella/enzimologia , Sequência de Bases , DNA Bacteriano , Dados de Sequência Molecular , Shewanella/genética
11.
Pest Manag Sci ; 57(5): 437-42, 2001 May.
Artigo em Inglês | MEDLINE | ID: mdl-11374161

RESUMO

Osmotic-sensitive (os-1) mutant alleles in Neurospora crassa exhibit resistance to dicarboximides, aromatic hydrocarbons and phenylpyrroles. We have previously reported that the os-1 mutants can be classified into two groups based on their resistance to fungicides and osmotic stress: type I, which are highly resistant to iprodione and fludioxonil but moderately sensitive to osmotic stress, and type II, which are highly sensitive to osmotic stress but moderately resistant to fungicides. To explain the mechanism of resistance to these fungicides, we cloned and sequenced the mutant os-1 genes that encode putative osmo-sensing histidine kinase. Within the os-1 gene product (Os1p), the type I strains, NM233t and Y256M209, carried a stop codon at amino acid position 308 and a frameshift at amino acid position 294, respectively. These mutation sites were located on the upstream of histidine kinase and the response regulator domains of Os1p, strongly suggesting that type I strains are null mutants. The null mutants, NM233t and Y256M209, were highly resistant to iprodione and fludioxonil; thus Os1p is essential for these fungicides to express their antifungal activity. The amino acid changes in Os1p, 625Pro from Leu, 578Val from Ala, and 580Arg from Gly were found in the type II strains, M16, M155-1 and P5990, respectively. Os1p is novel in having six tandem repeats of 90 amino acids in the N terminal. Each amino acid change of the type II strains was located on the fifth unit of six tandem repeats. Type II strains with single amino acid changes were more sensitive to osmotic stress than the null mutants (type I), indicating that the amino acid repeats of Os1p were responsible for an important function in osmo-regulation.


Assuntos
Aminoimidazol Carboxamida/análogos & derivados , Dioxóis/farmacologia , Proteínas Fúngicas/genética , Fungicidas Industriais/farmacologia , Genes Fúngicos , Hidantoínas , Mutação , Neurospora crassa/genética , Proteínas Quinases/genética , Pirróis/farmacologia , Sequência de Aminoácidos , Aminoimidazol Carboxamida/farmacologia , Clonagem Molecular , DNA Fúngico/genética , Resistência Microbiana a Medicamentos/genética , Dados de Sequência Molecular , Neurospora crassa/enzimologia , Neurospora crassa/fisiologia , Pressão Osmótica , Reação em Cadeia da Polimerase , Proteínas Quinases/metabolismo , Alinhamento de Sequência , Transdução de Sinais/genética
12.
Pediatr Surg Int ; 17(2-3): 104-7, 2001 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-11315264

RESUMO

Transforming growth factor-beta 1 (TGF beta-1) is an important mediator of liver-cell proliferation and replication that is implicated in hepatic fibrosis (HF). Hepatic stellate cells (HSC) are activated by TGF beta-1 and are the main precursor cells involved in fibrogenesis. The correlation between serum TGF beta-1, activated HSC in liver-biopsy specimens, and liver biochemistry was investigated to determine the value of TGF beta-1 as an indicator of clinical status in postoperative biliary atresia (BA) patients. Thirty-two postoperative BA patients (mean age 11.2 +/- 2.8 years) and 13 normal controls (mean age 10.3 +/- 3.7 years) were studied. Based on average liver function test (LFT) results over a 3-month period immediately prior to this study, the BA patients were divided into group I (anicteric, normal LFT; n = 10); group II (anicteric, elevated liver transaminases; n = 12), and group III (jaundiced end-stage liver fibrosis awaiting liver transplantation; n = 10). Serum TGF beta-1 was determined using ELISA. Liver-biopsy specimens were examined with antibody against TGF beta-1 and alpha-smooth muscle actin (SMA) antibody for detection of activated HSC. Serum TGF beta-1 was significantly higher in groups I (11.4 +/- 3.7 ng/ml; P < 0.01) and II (23.3 +/- 11.3 ng/ml; P < 0.001) than in group III (3.0 +/- 1.5 ng/ml) and controls (4.5 +/- 2.5 ng/ml) despite normal LFT in group I. The 3 subjects with the highest serum TGF beta-1 in group II had bile lakes. Biopsies from groups I and II were strongly positive for TGF beta-1 in hepatocytes and Kupffer cells and for activated HSC detected by SMA compared with group III and controls. Because serum TGF beta-1 and activated HSC are only present during active fibrosis, we conclude that there is progressive fibrogenesis even in seemingly normal postoperative BA patients. In particular, bile lakes should be regarded as a key sign of progressive HF, the presence of which should be regarded with extreme caution. We suggest that serum TGF beta-1 could be used as an accurate indicator of progressive fibrogenesis in postoperative BA patients.


Assuntos
Atresia Biliar/cirurgia , Cirrose Hepática/patologia , Complicações Pós-Operatórias/patologia , Fator de Crescimento Transformador beta/sangue , Adolescente , Atresia Biliar/patologia , Biópsia , Criança , Feminino , Seguimentos , Humanos , Fígado/patologia , Testes de Função Hepática , Masculino
13.
Biosci Biotechnol Biochem ; 65(2): 438-41, 2001 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-11302184

RESUMO

A thermostable beta-glucosidase from Thermus sp. Z-1 that not only hydrolyzes beta-glucosides but also beta-galactosides was shown to efficiently produce oligosaccharides during hydrolysis of lactose. The yield of oligosaccharides was more than 40% for 0.88 M lactose solution at 70 degrees C at pH 7.0. The major product was a trisaccharide, 3'-galactosyllactose, formed by a galactosyltransfer reaction.


Assuntos
Lactose/metabolismo , Oligossacarídeos/biossíntese , Thermus/enzimologia , beta-Glucosidase/metabolismo , Estabilidade Enzimática , Escherichia coli/genética , Temperatura Alta , Concentração de Íons de Hidrogênio , Cinética , Espectroscopia de Ressonância Magnética , Proteínas Recombinantes/genética , Proteínas Recombinantes/isolamento & purificação , Proteínas Recombinantes/metabolismo , Especificidade por Substrato , Thermus/genética , Trissacarídeos/biossíntese , beta-Glucosidase/genética , beta-Glucosidase/isolamento & purificação
14.
J Antibiot (Tokyo) ; 54(1): 10-6, 2001 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-11269706

RESUMO

A new cell growth inhibitor, curvularol, was isolated from the fermentation broth of Curvularia sp. RK97-F166. Curvularol showed no antibacterial activity, and very weak antifungal activity. However, curvularol inhibited the cell cycle progression of normal rat kidney (NRK) cells in G1 phase at 150 ng/ml. Curvularol induced the morphological reversion of srcts-transformed NRK cells at 100 ng/ml, and inhibited protein synthesis same as cycloheximide.


Assuntos
Ciclo Celular/efeitos dos fármacos , Fungos Mitospóricos/metabolismo , Inibidores da Síntese de Proteínas/isolamento & purificação , Inibidores da Síntese de Proteínas/farmacologia , Tricotecenos/isolamento & purificação , Tricotecenos/farmacologia , Animais , Linhagem Celular , DNA/biossíntese , DNA/efeitos dos fármacos , Fermentação , Humanos , Concentração Inibidora 50 , Testes de Sensibilidade Microbiana , Fungos Mitospóricos/classificação , Estrutura Molecular , Inibidores da Síntese de Proteínas/metabolismo , RNA/biossíntese , RNA/efeitos dos fármacos , Ratos , Tricotecenos/metabolismo , Quinases da Família src/efeitos dos fármacos , Quinases da Família src/genética , Quinases da Família src/metabolismo
15.
Biosci Biotechnol Biochem ; 65(1): 190-3, 2001 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-11272828

RESUMO

The gene encoding the principal a factor (rpoD) of the piezophilic bacterium Shewanella violacea was cloned and sequenced. The rpoD gene was found to encode a polypeptide consisting of 614 amino acid residues, showing 75.6 and 64.3% identity to those of Escherichia coli and Pseudomonas putida, respectively. Comparison with E. coli sigma70 and P. putida sigma70 showed that significant similarity exists in four conserved regions known to be required for promoter recognition and core binding. Using an expression plasmid harboring the rpoD gene, the S. violacea sigma70 factor was overexpressed in E. coli and successfully purified to near homogeneity.


Assuntos
Proteínas de Bactérias/genética , Proteínas de Ligação a DNA/genética , RNA Polimerases Dirigidas por DNA/genética , Shewanella/genética , Fator sigma/genética , Sequência de Aminoácidos , Proteínas de Bactérias/química , Proteínas de Bactérias/isolamento & purificação , Sequência de Bases , Proteínas de Ligação a DNA/química , Proteínas de Ligação a DNA/isolamento & purificação , RNA Polimerases Dirigidas por DNA/química , RNA Polimerases Dirigidas por DNA/isolamento & purificação , Eletroforese em Gel de Poliacrilamida , Escherichia coli/genética , Escherichia coli/metabolismo , Genes Bacterianos , Dados de Sequência Molecular , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Alinhamento de Sequência , Fator sigma/química , Fator sigma/isolamento & purificação
16.
Biosci Biotechnol Biochem ; 65(1): 205-8, 2001 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-11272832

RESUMO

Probenazole (PBZ) induces non-race specific resistance in rice plants against rice blast fungus and PBZ1 was identified as a PBZ-inducible gene from rice. The induction of PBZ1 expression in suspension-cultured rice cells was investigated. Northern blot analysis indicated that PBZ1 was induced by PBZ in a dose-dependent manner. Enzyme-linked immunosorbent assay (ELISA) showed a dose and time-dependent accumulation of PBZ1 protein. Both mRNA and protein analysis showed that PBZ1 was not induced by salicylic acid or an active metabolite, 1,2-benzisothiazole-1,1-dioxide.


Assuntos
Regulação da Expressão Gênica de Plantas , Oryza/genética , Proteínas de Plantas/biossíntese , Tiazóis/farmacologia , Linhagem Celular , Oryza/química , Oryza/efeitos dos fármacos , Proteínas de Plantas/genética , Ácido Salicílico/farmacologia
17.
Jpn J Pharmacol ; 85(1): 70-4, 2001 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-11243577

RESUMO

To clarify the mechanism for the severe emesis concomitant with intensive chemotherapy, we investigated the effects of 5-HT3- and 5-HT4-receptor antagonists on the emesis induced by the high-dose of cisplatin in Suncus murinus. The emesis induced by 50 mg/kg of cisplatin was reduced by the oral pretreatment with tropisetron, which is known as a 5-HT3- and 5-HT4-receptor dual antagonist in vitro, with the ID50 value of 0.52 mg/kg. On the contrary, granisetron, a selective 5-HT3-receptor antagonist, did not markedly inhibit the emesis at up to 30 mg/kg. Moreover, GR125487, a selective 5-HT4-receptor antagonist, did not inhibit the emesis. However, co-administration of GR125487 and granisetron significantly reduced the number of emetic episodes. The study of the co-administration of GR125487 with tropisetron showed that GR125487 did not further enhance the inhibitory effect of tropisetron alone, suggesting that the anti-emetic effect of tropisetron is mediated via the blockade of both 5-HT3 and 5-HT4 receptors. These results suggest that both the 5-HT3 and 5-HT4 receptors are involved in the emesis induced by the high-dose of cisplatin in Suncus murinus.


Assuntos
Antieméticos/farmacologia , Antineoplásicos/efeitos adversos , Cisplatino/efeitos adversos , Receptores de Serotonina/efeitos dos fármacos , Antagonistas da Serotonina/farmacologia , Vômito/fisiopatologia , Animais , Granisetron/farmacologia , Indóis/farmacologia , Masculino , Ondansetron/farmacologia , Receptores de Serotonina/fisiologia , Receptores 5-HT3 de Serotonina , Receptores 5-HT4 de Serotonina , Musaranhos , Sulfonamidas/farmacologia , Tropizetrona , Vômito/induzido quimicamente , Vômito/prevenção & controle
18.
Trends Biotechnol ; 19(3): 102-8, 2001 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-11179803

RESUMO

Microorganisms that prefer high-pressure conditions are termed piezophiles (previously termed barophiles). The molecular basis of piezophily is now being investigated extensively focusing on aspects of gene regulation and the function of certain proteins in deep-sea isolates. Little attention has been paid, however, to the potential biotechnological applications of piezophiles compared with other extremophiles. Based on the fundamental knowledge available, we will try to answer the following questions: How can we exploit the biotechnological potential of piezophiles? What can be understood by the application of high-pressure in biological systems?


Assuntos
Biotecnologia/tendências , Divisão Celular , Enzimas/metabolismo , Regulação da Expressão Gênica , Pressão Hidrostática , Microbiologia , Proteínas/química , Shewanella/genética , Shewanella/crescimento & desenvolvimento , Shewanella/metabolismo
19.
Mol Cell Biochem ; 216(1-2): 111-9, 2001 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-11216855

RESUMO

We investigated changes in cytoplasmic Ca2+ concentration ([Ca2+]i) and in left ventricular contractility during sustained ischemia and reperfusion in isolated beating rat hearts. Hearts from male Sprague-Dawley rats were perfused retrogradely and were loaded with 4 microM fura-2. Low-flow global ischemia was induced by reducing perfusion flow to 10% and by electric pacing. The hearts were exposed to ischemia for 10 min or 30 min and then reperfused. [Ca2+]i was measured by monitoring the ratio of 500 nm fluorescence excited at 340 and 380 nm while simultaneously measuring left ventricular pressure (LVP). To determine diastolic [Ca2+]i, background autofluorescence was subtracted. LVP rapidly decreased from 82.3+/-8.2 to 17.1+/-2.9 mmHg , whereas the amplitude of the Ca2+ transient did not change significantly during the first 1 min of ischemia. After 10 min of ischemia, the amplitude decreased to 60.8+/-10.6% (p < 0.05) and diastolic [Ca2+]i increased by 26.3+/-2.9% (p < 0.001) compared with the pre-ischemic value (n = 8). When the hearts were reperfused after 10 min of ischemia, the amplitude of the Ca2+ transient and LVP recovered to 79.0+/-7.2% and 73.2+/-7.5 mmHg, respectively. Whereas diastolic [Ca2+]i decreased to the preischemic value. In the hearts exposed to 30 min of ischemia (n = 10), diastolic [Ca2+]i increased even further by 32.7+/-5.3% at the end of ischemia and continued increasing during the 10 min of reperfusion by 42.6+/-15.6%. Six of 10 hearts developed ventricular fibrillation (VF) and intracellular Ca2+ overload after reperfusion. Recovery of LVP after reperfusion was significantly smaller in the hearts exposed to 30 min of ischemia than in the hearts exposed to 10 min of ischemia (58.9+/-11.7 vs. 97.2+/-3.0% of pre-ischemic value, p < 0.05). Diastolic [Ca2+]i also increased under hypoxic conditions (N2 bubbling) in this model. These results suggest that increases in diastolic [Ca2+]i might play an important role in myocardial contractile dysfunction and viability in ischemia-reperfusion injury.


Assuntos
Cálcio/metabolismo , Coração/fisiologia , Contração Miocárdica , Isquemia Miocárdica , Miocárdio/metabolismo , Traumatismo por Reperfusão , Animais , Diástole , Corantes Fluorescentes/farmacologia , Fura-2/farmacologia , Hipóxia , Masculino , Perfusão , Ratos , Ratos Sprague-Dawley , Espectrometria de Fluorescência , Fatores de Tempo , Função Ventricular Esquerda
20.
Antonie Van Leeuwenhoek ; 80(3-4): 317-23, 2001 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-11827218

RESUMO

A new species of the genus Rhodotorula was isolated from a tubeworm (Lamellibrachia sp.) collected at a depth of 1156 m in Sagami Bay, Japan. Strain SY-89 had physiological properties quite similar to R. aurantiaca. Two phylogenetic trees, one based on internal transcribed spacer (ITS) regions and 5.8S rDNA sequences and the other based on the D1/D2 region of the large subunit (26S) rDNA sequences, united strain SY-89 to the type strain of Sakaguchia dacryoides through a considerable evolutionary distance. Strain SY-89 was differentiated from S. dacryoides by the G+C content of the nuclear DNA and differences in the ability to utilize specific carbon and nitrogen compounds. The low complementarity of strain SY-89 DNA to that of the type strain of S. dacryoides confirmed that this strain was genetically unrelated to previously known species. The tubeworm isolates are described as R. lamellibrachii sp. nov. The type strain of R. lamellibrachii is strain SY-89 (= JCM 10907). R. lamellibrachii formed a cluster with Erythrobasidium hasegawianum, R. lactosa, S. dacryoides and Sporobolomyces elongatus on the ITS and 5.8S rDNA phylogenetic tree. These five species shared a signature sequence in 26S rDNA, although this relationship was not supported by phylogeny based on the D1/D2 region of 26S rDNA.


Assuntos
Poliquetos/microbiologia , Rhodotorula/classificação , Rhodotorula/isolamento & purificação , Animais , DNA Fúngico/genética , DNA Ribossômico/genética , Japão , Biologia Marinha , Filogenia , RNA Fúngico/genética , RNA Ribossômico/genética , RNA Ribossômico 5,8S/genética , Rhodotorula/genética , Especificidade da Espécie
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