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1.
Clin Nutr Res ; 6(2): 122-129, 2017 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-28503508

RESUMO

The aim of this study was to investigate the association between macronutrient composition and metabolic syndrome (MetS) incidence in Korean adults. Data were obtained from a cohort of 10,030 members aged 40 to 69 years who were enrolled from the 2 cities (Ansung and Ansan) between 2001 and 2002 to participate in the Korean Genome Epidemiology Study. Of these members, 5,565 participants, who were free of MetS and reported no diagnosis of cardiovascular disease at baseline, were included in this study. MetS was defined using the criteria of the National Cholesterol Education Program-Adult Treatment Panel III and Asia-Pacific criteria for waist circumference. MetS incidence rate were identified during a 2-year follow-up period. Baseline dietary information was obtained using a semi-quantitative food frequency questionnaire. Multivariate logistic regression analysis was used to evaluate the association between the quartiles of percentages of total calorie from macronutrients consumed and MetS incidence. In analyses, baseline information, including age, sex, body mass index, income status, educational status, smoking status, alcohol drinking status, and physical activity level was considered as confounding variables. Participants with the second quartile of the percentages of carbohydrate calorie (67%-70%) had a 23% reduced odds ratio (95% confidence interval, 0.61-0.97) for MetS incidence compared with those with the fourth quartile after adjusting for confounding variables. The findings suggest that middle aged or elderly Korean adults who consume approximately 67%-70% of calorie from carbohydrate have a reduced risk of MetS.

2.
Ultrasound Obstet Gynecol ; 49(3): 315-321, 2017 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-27546354

RESUMO

OBJECTIVE: Randomized controlled trials (RCTs) have recently compared intramuscular 17α-hydroxyprogesterone caproate (17-OHPC) with vaginal progesterone for reducing the risk of spontaneous preterm birth (SPTB) in singleton gestations with prior SPTB. The aim of this systematic review and meta-analysis was to evaluate the efficacy of vaginal progesterone compared with 17-OHPC in prevention of SPTB in singleton gestations with prior SPTB. METHODS: Searches of electronic databases were performed to identify all RCTs of asymptomatic singleton gestations with prior SPTB that were randomized to prophylactic treatment with either vaginal progesterone (intervention group) or intramuscular 17-OHPC (comparison group). No restrictions for language or geographic location were applied. The primary outcome was SPTB < 34 weeks. Secondary outcomes were SPTB < 37 weeks, < 32 weeks, < 28 weeks and < 24 weeks, maternal adverse drug reaction and neonatal outcomes. The summary measures were reported as relative risk (RR) with 95% CI. Risk of bias for each included study was assessed. RESULTS: Three RCTs (680 women) were included. The mean gestational age at randomization was about 16 weeks. Women were given progesterone until 36 weeks or delivery. Regarding vaginal progesterone, one study used 90 mg gel daily, one used 100 mg suppository daily and one used 200 mg suppository daily. All included RCTs used 250 mg intramuscular 17-OHPC weekly in the comparison group. Women who received vaginal progesterone had significantly lower rates of SPTB < 34 weeks (17.5% vs 25.0%; RR, 0.71 (95% CI, 0.53-0.95); low quality of evidence) and < 32 weeks (8.9% vs 14.5%; RR, 0.62 (95% CI, 0.40-0.94); low quality of evidence) compared with women who received 17-OHPC. There were no significant differences in the rates of SPTB < 37 weeks, < 28 weeks and < 24 weeks. The rate of women who reported adverse drug reactions was significantly lower in the vaginal progesterone group compared with the 17-OHPC group (7.1% vs 13.2%; RR, 0.53 (95% CI, 0.31-0.91); very low quality of evidence). Regarding neonatal outcomes, vaginal progesterone was associated with a lower rate of neonatal intensive care unit admission compared with 17-OHPC (18.7% vs 23.5%; RR, 0.63 (95% CI, 0.47-0.83); low quality of evidence). For the comparison of 17-OHPC vs vaginal progesterone, the quality of evidence was downgraded for all outcomes by at least one degree due to imprecision (the optimal information size was not reached) and by at least one degree due to indirectness (different interventions). CONCLUSIONS: Daily vaginal progesterone (either suppository or gel) started at about 16 weeks' gestation is a reasonable, if not better, alternative to weekly 17-OHPC injection for prevention of SPTB in women with singleton gestations and prior SPTB. However, the quality level of the summary estimates was low or very low as assessed by GRADE, indicating that the true effect may be, or is likely to be, substantially different from the estimate of the effect. Copyright © 2016 ISUOG. Published by John Wiley & Sons Ltd. COMPARACIÓN ENTRE LA PROGESTERONA VAGINAL Y EL 17Α-HIDROXIPROGESTERONA CAPROATO INTRAMUSCULAR PARA LA PREVENCIÓN DEL PARTO PRETÉRMINO ESPONTÁNEO RECURRENTE EN EMBARAZOS CON FETO ÚNICO: REVISIÓN SISTEMÁTICA Y METAANÁLISIS DE ENSAYOS CONTROLADOS ALEATORIOS: RESUMEN OBJETIVO: Recientemente se han realizado varios ensayos controlados aleatorios (ECA) que comparaban el caproato de 17α-hidroxiprogesterona (17-OHPC, por sus siglas en inglés) por vía intramuscular con la progesterona por vía vaginal para la reducción del riesgo de parto pretérmino espontáneo (PPTE) en embarazos con feto único de gestantes con historial de PPTE. El objetivo de esta revisión sistemática y metaanálisis fue evaluar la eficacia de la progesterona vaginal en comparación con la 17-OHPC en la prevención de embarazos con feto único de gestantes con historial de PPTE. MÉTODOS: Se realizaron búsquedas en bases de datos electrónicas para identificar todos los ECA con embarazos de feto único asintomáticos con historial de PPTE antes de ser asignados al azar a un tratamiento profiláctico, ya fuera con progesterona vaginal (grupo de intervención) o con 17-OHPC intramuscular (grupo de control). No se aplicaron restricciones respecto al idioma o la ubicación geográfica. El resultado primario fue PPTE < 34 semanas. Los resultados secundarios fueron PPTE <37 semanas, < 32 semanas, < 28 semanas y < 24 semanas, la reacción materna adversa al fármaco y los resultados neonatales. Las medidas del resumen se reportaron como riesgo relativo (RR) con IC del 95%. Para cada estudio incluido se evaluó el riesgo de sesgo. RESULTADOS: Se incluyeron tres ECA (680 mujeres). La media de la edad gestacional en el momento de la aleatorización fue de 16 semanas. A las mujeres se les administró progesterona hasta la semana 36 o hasta el parto. Con respecto a la progesterona vaginal, un estudio utilizó gel de 90 mg diariamente, otro utilizó un supositorio diario de 100 mg y el otro utilizó un supositorio diario de 200 mg. Todos los ECA incluidos en el grupo de comparación utilizaron 250 mg semanales de 17-OHPC por vía intramuscular. Las mujeres que recibieron progesterona vaginal tuvieron tasas significativamente más bajas de PPTE < 34 semanas (17,5% vs. 25,0%; RR 0,71 (IC 95%, 0,53-0,95); calidad de la evidencia baja) y < 32 semanas (8,9% vs. 14,5%; RR 0,62 (IC 95%, 0,40-0,94); calidad de evidencia baja), en comparación con las mujeres que recibieron 17-OHPC. No hubo diferencias significativas en las tasas de PPTE < 37 semanas, < 28 semanas y < 24 semanas. La tasa de mujeres que reportaron reacciones adversas a los medicamentos fue significativamente menor en el grupo de progesterona vaginal en comparación con el grupo de 17-OHPC (7,1% vs. 13,2%; RR 0,53 (IC 95%, 0,31-0,91); calidad de la evidencia muy baja). En cuanto a los resultados neonatales, la progesterona vaginal se asoció a una menor tasa de admisiones en la unidad neonatal de cuidados intensivos en comparación con la 17-OHPC (18,7% vs. 23,5%; RR 0,63 (IC 95%, 0,47-0,83); calidad de evidencia baja). Para la comparación del 17-OHPC con la progesterona vaginal se rebajó la calidad de las pruebas para todos los resultados en al menos un grado debido a imprecisiones (no se alcanzó el tamaño óptimo de la información) y en al menos un grado debido al carácter indirecto de los estudios (diferentes intervenciones). CONCLUSIONES: La progesterona vaginal administrada diariamente (ya fuera como supositorio o como gel) desde la semana 16 de gestación es una alternativa razonable, si no mejor, a una inyección semanal de 17-OHPC para la prevención de PPTE en mujeres con embarazos de feto único e historial de PPTE. Sin embargo, el nivel de calidad de las estimaciones del resumen fue bajo o muy bajo según lo evaluado por GRADE, lo que indica que el verdadero efecto puede ser, o es probable que sea, sustancialmente diferente de la estimación del efecto. 17Α-:META: : (randomized controlled trials,RCTs)(spontaneous preterm birth,SPTB)17α-(intramuscular 17α-hydroxyprogesterone caproate,17-OHPC)SPTB。metaSPTB17-OHPCSPTB。 : ,SPTBRCTs,RCTs()17-OHPC()。。34SPTB。37、32、2824SPTB,。(relative risk,RR)95%CI。。 : 3RCTs(680)。16。,36。,90 mg,100 mg,200 mg。,RCTs250 mg 17-OHPC。17-OHPC,34 [17.5%25.0%;RR,0.71(95% CI,0.53 ~ 0.95);]32[8.9%14.5%;RR,0.62(95% CI,0.40 ~ 0.94);]SPTB。37、2824SPTB。17-OHPC,[7.1%13.2%;RR,0.53(95% CI,0.31 ~ 0.91);]。,17-OHPC,[18.7%23.5%;RR,0.63(95% CI,0.47 ~ 0.83);]。17-OHPC,(),()。 : SPTBSPTB,16()17-OHPC,。,GRADE,,。.


Assuntos
Hidroxiprogesteronas/administração & dosagem , Nascimento Prematuro/prevenção & controle , Progesterona/administração & dosagem , Caproato de 17 alfa-Hidroxiprogesterona , Administração Intravaginal , Esquema de Medicação , Feminino , Humanos , Hidroxiprogesteronas/uso terapêutico , Injeções Intramusculares , Unidades de Terapia Intensiva , Admissão do Paciente/estatística & dados numéricos , Gravidez , Progesterona/uso terapêutico , Ensaios Clínicos Controlados Aleatórios como Assunto
3.
J Viral Hepat ; 22(11): 948-56, 2015 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-25817821

RESUMO

We investigated an outbreak of jaundice in urban Bangladesh in 2010 to examine the cause and risk factors and assess the diagnostic utility of commercial assays. We classified municipal residents reporting jaundice during the preceding 4 weeks as probable hepatitis E cases and their neighbours without jaundice in the previous 6 months as probable controls. We tested the sera collected from probable cases and probable controls for IgM anti-hepatitis E virus (HEV), and the IgM-negative sera for IgG anti-HEV using a commercial assay locally. We retested the IgM-positive sera for both IgM and IgG anti-HEV using another assay at the Centre for Disease Control and Prevention (CDC), USA. Probable cases positive for IgM anti-HEV were confirmed cases; probable controls negative for both IgM and IgG anti-HEV were confirmed controls. We explored the local water supply and sanitation infrastructure and tested for bacterial concentration of water samples. Probable cases were more likely than probable controls to drink tap water (adjusted odds ratio: 3.4; 95% CI: 1.2-9.2). Fifty-eight percentage (36/62) of the case sera were IgM anti-HEV positive; and 75% of the IgM-positive samples were confirmed positive on retesting with another assay at CDC. Compared to confirmed controls, cases confirmed using either or both assays also identified drinking tap water as the risk factor. Two tap water samples had detectable thermotolerant coliforms. Research exploring decentralized water treatment technologies for sustainable safe water might prevent HEV transmission in resource-poor cities. Detection of serological markers in a majority of probable cases implied that available diagnostic assays could adequately identify HEV infection during outbreaks.


Assuntos
Surtos de Doenças , Hepatite E/epidemiologia , Icterícia/etiologia , População Urbana , Adolescente , Adulto , Bangladesh/epidemiologia , Estudos de Casos e Controles , Cidades/epidemiologia , Feminino , Anticorpos Anti-Hepatite/sangue , Hepatite E/diagnóstico , Hepatite E/patologia , Humanos , Imunoensaio , Imunoglobulina G/sangue , Imunoglobulina M/sangue , Icterícia/diagnóstico , Icterícia/patologia , Masculino , Microbiologia da Água , Adulto Jovem
4.
Mymensingh Med J ; 23(2): 341-4, 2014 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-24858164

RESUMO

More than ninety percent of bladder neoplasm is Transitional Cell Carcinoma (TCC). About 85% of patients present with symptom of painless haematuria. However haematuria is quite often intermittent so that a negative result has little meaning in ruling out the presence of bladder cancer. The present study was conducted to observe the natural history of different clinicopathologic stages of bladder cancer after transurethral resection bladder tumor (TURBT), either intravescical chemotherapy (IVC) or immunotherapy with BCG and of other forms of therapy like radio-chemotherapy (RCT) and radical surgery in few cases. A consecutive series of 52 cases with different grades and stages of bladder cancer patients were studied. Those who were noncompliant with surgery were sent for RCT. The age range of the patients was 34 to 75 with mean 53 year. Macroscopic haematuria, flank pain, microscopic haematuria, bladder irritability was found in 43(82.7%), 06(11.5%), 03(5.8%), 32(61.5%) cases respectively and coincidental ureteral TCC with hydronephrosis was found in 04(7.7%) cases for whom radical nephreureterectomy and cystectomy with ileal conduit was done. Amongst the other muscle invasive tumor only three cases were compliant to do radical surgery; rest were advised to consult with oncologist for possible RCT. Superficial bladder cancer was 19(36.5%) and invasive bladder cancer was 33(63.5%). TURBT and IVT were offered for all superficial bladder tumors. Of them 06(31.5%) patients showed recurrence during the study period. More recurrences occur in IVC group (35.7%) than immunotherapy with BCG group (20%) Re-TURBT and stage wise treatment was offered to them. All recurrent cases were G-3 tumor and were multifocal. Recurrence rate is about 30%. Screening program and structured referral system should be developed to have early diagnosis for prompt treatment and best prognosis.


Assuntos
Carcinoma de Células de Transição/patologia , Carcinoma de Células de Transição/terapia , Neoplasias da Bexiga Urinária/patologia , Neoplasias da Bexiga Urinária/terapia , Adulto , Idoso , Estudos de Coortes , Terapia Combinada/métodos , Feminino , Hematúria/etiologia , Hematúria/patologia , Hematúria/terapia , Humanos , Masculino , Pessoa de Meia-Idade , Resultado do Tratamento
8.
J Appl Microbiol ; 93(5): 800-9, 2002.
Artigo em Inglês | MEDLINE | ID: mdl-12392526

RESUMO

AIMS: Effective household hygiene measures require that sources of bacterial contamination and the places to which contamination spreads be carefully identified. Therefore, a study was performed to examine the distribution of microorganisms throughout ordinary households in Japan, which has its own unique customs of daily life and food preparation. METHODS AND RESULTS: Using the stamping method, samples were taken from 100 different places and items in each of 86 households. This study found kitchens/dining rooms to have the greatest level of microbial contamination and bathrooms, the next highest level. Toilets (water closets) were found to have an unexpectedly low level of bacterial contamination. The largest bacterial counts were found on items such as drain traps, dish-washing sponges, counter towels, sinks, dish-washing tubs, and bathroom sponges. CONCLUSIONS: It is necessary to carefully identify both the items that can become instruments for spreading bacterial contamination and the places that easily become subject to secondary contamination, and then to take timely and effective disinfection/sanitizing measures. SIGNIFICANCE AND IMPACT OF THE STUDY: The data gathered in this study will be very valuable for anticipating the pathways over which bacteria are transported and prioritizing disinfection targets, to make effective disinfection possible.


Assuntos
Bactérias/isolamento & purificação , Higiene , Pseudomonas aeruginosa/isolamento & purificação , Atividades Cotidianas , Criança , Culinária , Cultura , Enterobacteriaceae/crescimento & desenvolvimento , Enterobacteriaceae/isolamento & purificação , Escherichia coli/crescimento & desenvolvimento , Escherichia coli/isolamento & purificação , Feminino , Contaminação de Alimentos/análise , Fungos/crescimento & desenvolvimento , Fungos/isolamento & purificação , Habitação , Humanos , Japão , Masculino , Pseudomonas aeruginosa/crescimento & desenvolvimento , Saneamento , Staphylococcus aureus/crescimento & desenvolvimento , Staphylococcus aureus/isolamento & purificação , Toaletes
10.
Int J Environ Health Res ; 12(1): 41-52, 2002 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-11970814

RESUMO

The bacterial contamination of Japanese homes and the attitudes of Japanese people toward sanitation were studied. By taking bacterial counts of approximately 90 places each in five homes, this study found kitchens to have the greatest degree of bacterial contamination, followed by bathing rooms. Toilets had less bacterial contamination than was expected. While concern about bacteria on highly contaminated items such as sponges, towels for wiping counters, and other reservoirs/disseminators was common, there was a relative lack of concern regarding contact surfaces such as dining tables. It is believed that an in-depth study of bacterial contamination in the home and concern about it would lead to the promotion of greater public understanding of home sanitation and help to facilitate the provision of useful information and products to the public.


Assuntos
Escherichia coli/isolamento & purificação , Contaminação de Alimentos , Pseudomonas aeruginosa/isolamento & purificação , Saneamento , Staphylococcus aureus/isolamento & purificação , Toaletes , Atividades Cotidianas , Culinária , Escherichia coli/patogenicidade , Habitação , Humanos , Higiene , Japão , Pseudomonas aeruginosa/patogenicidade , Staphylococcus aureus/patogenicidade
11.
Nihon Ishinkin Gakkai Zasshi ; 42(4): 253-8, 2001.
Artigo em Japonês | MEDLINE | ID: mdl-11704756

RESUMO

The infection rate, causative dermatophytes, and dermatophyte dissemination of tinea pedis in young healthy Japanese were studied by direct microscopic examination, slant cultures, and foot-press culture method. Questionnaires on subjective symptoms and treatments were also distributed. Among fifty-eight medical students with a mean age of 23.9 years, thirteen (22.4%) showed positive by direct microscopic examination and T. mentagrophytes was more dominant than T. rubrum by slant cultures. In one hundred and sixteen student feet, twelve were infected and disseminating dermatophytes, four were infected but not disseminating, three were not infected but adhering dermatophytes. The infection rate of tinea pedis was thus 24.1%. Among thirty-seven research workers (mean age: 34.8), twenty-one (56.8%) showed positive by direct microscopic examination. All the dermatophytes isolated by slant cultures were T. mentagrophytes. In seventy-four feet, twenty-nine were infected and disseminating, ten were infected but not disseminating, and three were adhering dermatophytes. The infection rate was 64.9%. Among thirty-one females wearing boots (mean age: 21.0), seven (22.6%) were infected and T. mentagrophytes was more dominant. In sixty-two feet, eight were infected and disseminating, one was infected but not disseminating, and five were adhering dermatophytes. The infection rate of tinea pedis was quite high and T.r/T.m rate was low in the three groups. Most of the patients had had no treatment and were disseminating dermatophytes.


Assuntos
, Sapatos/efeitos adversos , Pele/microbiologia , Tinha dos Pés/epidemiologia , Adulto , Feminino , Humanos , Japão/epidemiologia , Masculino , Tinha dos Pés/microbiologia , Trichophyton/isolamento & purificação , Trichophyton/patogenicidade
12.
J Vet Med Sci ; 63(12): 1297-302, 2001 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-11789607

RESUMO

The structure of isotype-specific regions of classes 1, II, III, IVa and IVb of canine beta-tubulin was characterized by 3'-RACE and the expression of these isotypes in canine tissues was examined by ribonuclease protection assay (RPA). Furthermore, a malignant mammary tumor-derived osteosarcoma-like cell line was established and the altered expression of beta-tubulin isotypes in taxol-resistant sublines was analyzed. The deduced amino acid sequences in isotype-specific regions corresponding to classes I, II and IVb were identical to those of humans and mice, but those in classes III and IVa showed slight differences among species. RPA revealed that classes I and IVb were widely distributed, but classes II, III and IVa were restricted to the brain. Because RPA could clearly distinguish the expression of class IVa from that of class IVb, it was thought to be more useful than northern blot for analysis of beta-tubulin isotype expression. In vitro, taxol-resistant sublines displayed a significant increase in class IVa as compared with taxol-sensitive cells, suggesting that altered expression of class IVa was associated with taxol resistance in these cell lines.


Assuntos
Neoplasias Ósseas/veterinária , Doenças do Cão/metabolismo , Osteossarcoma/veterinária , Tubulina (Proteína)/classificação , Sequência de Aminoácidos , Animais , Antineoplásicos Fitogênicos/farmacologia , Neoplasias Ósseas/genética , Neoplasias Ósseas/metabolismo , Doenças do Cão/genética , Cães , Feminino , Regulação Neoplásica da Expressão Gênica , Masculino , Neoplasias Mamárias Animais/patologia , Camundongos , Camundongos SCID , Dados de Sequência Molecular , Osteossarcoma/genética , Osteossarcoma/metabolismo , Paclitaxel/farmacologia , Reação em Cadeia da Polimerase/veterinária , Isoformas de Proteínas/biossíntese , Isoformas de Proteínas/classificação , Isoformas de Proteínas/genética , Homologia de Sequência de Aminoácidos , Tubulina (Proteína)/biossíntese , Tubulina (Proteína)/genética , Células Tumorais Cultivadas
13.
Biosci Biotechnol Biochem ; 64(2): 399-404, 2000 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-10737199

RESUMO

Resting cells of a double mutant noted as KSM-MT66, derived from Rhodococcus sp. strain KSM-B-3 by UV irradiation, were found to cis-desaturate isopropyl hexadecanoate, yielding isopropyl cis-6-hexadecenoate. Addition of sodium glutamate (1.0%), Mg SO4 (2 mM), and thiamine (2 mM) increased the productivity of the unsaturated product in phosphate buffer. Optimal temperature and pH for the reaction were around 26 degrees C and 7, respectively. Under the optimized conditions, more than 50 g/l of isopropyl cis-6-hexadecenoate was produced after a 3-day incubation by resting cells of the mutant. Thus, cis-6-hexadecenoic acid, the main component of human sebaceous lipids, can be manufactured economically by the rhodococcal bioconversion.


Assuntos
Ésteres/metabolismo , Palmitatos/metabolismo , Rhodococcus/genética , Aminoácidos/metabolismo , Ácidos Graxos Monoinsaturados , Concentração de Íons de Hidrogênio , Metais/metabolismo , Mutação , Oxigênio/metabolismo , Rhodococcus/metabolismo , Temperatura , Vitaminas/metabolismo
14.
Appl Environ Microbiol ; 65(12): 5636-8, 1999 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-10584034

RESUMO

A mutant Rhodococcus strain lacking the ability to utilize 1-chlorohexadecane was found to cis-desaturate aliphatic compounds, such as 1-chlorohexadecane, n-hexadecane, and heptadecanonitrile, yielding corresponding products with a double bond mainly at the ninth carbon from the terminal methyl groups. A new oxidative pathway involving the cis-desaturation step was suggested for alkane utilization by Rhodococcus spp.


Assuntos
Hidrocarbonetos Clorados/metabolismo , Hidrocarbonetos/metabolismo , Rhodococcus/genética , Rhodococcus/metabolismo , Alcanos/metabolismo , Espectrometria de Massas , Mutação , Nitrilas/metabolismo , Espectroscopia de Infravermelho com Transformada de Fourier
15.
Extremophiles ; 2(3): 185-90, 1998 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-9783164

RESUMO

The cleaning power of detergents seems to have peaked; all detergents contain similar ingredients and are based on similar detergency mechanisms. To improve detergency, modern types of heavy-duty powder detergents and automatic dishwasher detergents usually contain one or more enzymes, such as protease, amylase, cellulase, and lipase. Alkaliphilic Bacillus strains are often good sources of alkaline extracellular enzymes, the properties of which fulfil the essential requirements for enzymes to be used in detergents. We have isolated numbers of alkaliphilic Bacillus that produce such alkaline detergent enzymes, including cellulase (CMCase), protease, alpha-amylase, and debranching enzymes, and have succeeded in large-scale industrial production of some of these enzymes. Here, we describe the enzymatic properties, genetics, and structures of the detergent enzymes that we have developed.


Assuntos
Bacillus/enzimologia , Detergentes/metabolismo , Enzimas/metabolismo , Sequência de Aminoácidos , Bacillus/genética , Proteínas de Bactérias/química , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Biotecnologia , Celulase/química , Celulase/genética , Celulase/metabolismo , Endopeptidases/química , Endopeptidases/genética , Endopeptidases/metabolismo , Enzimas/química , Enzimas/genética , Glicosídeo Hidrolases/química , Glicosídeo Hidrolases/genética , Glicosídeo Hidrolases/metabolismo , Concentração de Íons de Hidrogênio , Modelos Moleculares , Dados de Sequência Molecular , Filogenia , Homologia de Sequência de Aminoácidos
16.
Appl Environ Microbiol ; 64(9): 3282-9, 1998 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-9726872

RESUMO

A novel liquefying alpha-amylase (LAMY) was found in cultures of an alkaliphilic Bacillus isolate, KSM-1378. The specific activity of purified LAMY was approximately 5,000 U mg of protein-1, a value two- to fivefold greater between pH 5 and 10 than that of an industrial, thermostable Bacillus licheniformis enzyme. The enzyme had a pH optimum of 8.0 to 8.5 and displayed maximum activity at 55 degreesC. The molecular mass deduced from sodium dodecyl sulfate-polyacrylamide gel electrophoresis was approximately 53 kDa, and the apparent isoelectric point was around pH 9. This enzyme efficiently hydrolyzed various carbohydrates to yield maltotriose, maltopentaose, maltohexaose, and maltose as major end products after completion of the reaction. Maltooligosaccharides in the maltose-to-maltopentaose range were unhydrolyzable by the enzyme. The structural gene for LAMY contained a single open reading frame 1, 548 bp in length, corresponding to 516 amino acids that included a signal peptide of 31 amino acids. The calculated molecular mass of the extracellular mature enzyme was 55,391 Da. LAMY exhibited relatively low amino acid identity to other liquefying amylases, such as the enzymes from B. licheniformis (68.9%), Bacillus amyloliquefaciens (66.7%), and Bacillus stearothermophilus (68.6%). The four conserved regions, designated I, II, III, and IV, and the putative catalytic triad were found in the deduced amino acid sequence of LAMY. Essentially, the sequence of LAMY was consistent with the tertiary structures of reported amylolytic enzymes, which are composed of domains A, B, and C and which include the well-known (alpha/beta)8 barrel motif in domain A.


Assuntos
Bacillus/enzimologia , alfa-Amilases , Sequência de Aminoácidos , Bacillus/genética , Sequência de Bases , Cátions/farmacologia , Clonagem Molecular , DNA Bacteriano/análise , Estabilidade Enzimática , Concentração de Íons de Hidrogênio , Dados de Sequência Molecular , Reação em Cadeia da Polimerase , Alinhamento de Sequência , Especificidade por Substrato , Temperatura , alfa-Amilases/química , alfa-Amilases/genética , alfa-Amilases/isolamento & purificação , alfa-Amilases/metabolismo
17.
J Biol Chem ; 271(39): 24075-83, 1996 Sep 27.
Artigo em Inglês | MEDLINE | ID: mdl-8798645

RESUMO

An amylopullulanase from alkalophilic Bacillus sp. KSM-1378 hydrolyzes both alpha-1,6 linkages in pullulan and alpha-1,4 linkages in other polysaccharides, with maximum activity in each case at an alkaline pH, to generate oligosaccharides (Ara, K., Saeki, K., Igarashi, K., Takaiwa, M., Uemura, T., Hagihara, H., Kawai, S., and Ito, S. (1995) Biochim. Biophys. Acta 1243, 315-324). Here, we report the molecular cloning and sequencing of the gene for and the structure of this enzyme and show that its dual hydrolytic activities are associated with two independent active sites. The structural gene contained a single, long open reading frame of 5,814 base pairs, corresponding to 1,938 amino acids that included a signal peptide of 32 amino acids. The molecular mass of the extracellular mature enzyme (Glu33 through Leu1938) was calculated to be 211,450 Da, a value close to the 210 kDa determined for the amylopullulanase produced by Bacillus sp. KSM-1378. The amylase and the pullulanase domains were located in the amino-terminal half and in the carboxyl-terminal half of the enzyme, respectively, being separated by a tandem repeat of a sequence of 35 amino acids. Four regions, designated I, II, III, and IV, were highly conserved in each catalytic domain, and they included a putative catalytic triad Asp550-Glu579-Asp645 for the amylase activity and Asp1464-Glu1493-Asp1581 for the pullulanase activity. The purified enzyme was rotary shadowed at a low angle and observed by transmission electron microscopy; it appeared to be a "castanet-like" or "bent dumbbell-like" molecule with a diameter of approximately 25 nm.


Assuntos
Bacillus/enzimologia , Proteínas de Bactérias/química , Glucanos/metabolismo , Glicosídeo Hidrolases/química , Polissacarídeos/metabolismo , Sequência de Aminoácidos , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Sequência de Bases , Sítios de Ligação , Clonagem Molecular , Sequência Consenso , DNA Bacteriano/genética , Genes Bacterianos , Glicosídeo Hidrolases/genética , Glicosídeo Hidrolases/metabolismo , Dados de Sequência Molecular , Mapeamento por Restrição , Alinhamento de Sequência , Homologia de Sequência de Aminoácidos , Especificidade por Substrato
18.
Biosci Biotechnol Biochem ; 60(4): 634-9, 1996 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-8829530

RESUMO

An amylopullulanase (APase) from alkalophilic Bacillus sp. KSM-1378 hydrolyzes both alpha-1,6 linkages in pullulan and alpha-1,4 linkages in other polysaccharides, each maximally active at an alkaline pH, to generate oligosaccharides. We analyzed proteolytic fragments that were produced by exposing pure APase to various proteases, to identify its catalytic domain(s). The intact, pure 210-kDa APase was partially digested with papain for a short time, yielding simultaneously two smaller non-overlapping active fragments, designated amylose-hydrolyzing fragment (AHF114, 114 kDa) and pullulan-hydrolyzing fragment (PHF102, 102 kda). The two truncated protein fragments, each containing a single catalytic domain, were purified to homogeneity. The purified AHF114 and PHF102 had similar enzymatic properties to the amylase and pullulanase activities, respectively, of intact APase. The partial amino-terminal sequences of APase and AHF114 were both Glu-Thr-Gly-Asp-Lys-Arg-Ile-Glu-Phe-Ser-Tyr-Glu-Arg-Pro and that of PHF102 was Thr-Val-Pro-Leu-Ala-Leu-Val-Ser-Gly-Glu-Val-Leu-Ser-Asp-Lsy-Leu. These results were direct evidence that the alpha-1,6 and alpha-1,4 hydrolytic activities were associated with two different active sites in this novel enzyme. Our alkaline APase is obviously a "biheaded enzyme".


Assuntos
Bacillus/enzimologia , Proteínas de Bactérias/metabolismo , Glicosídeo Hidrolases/metabolismo , Papaína/metabolismo , Estrutura Terciária de Proteína , Sequência de Aminoácidos , Inibidores Enzimáticos/farmacologia , Estabilidade Enzimática , Concentração de Íons de Hidrogênio , Hidrólise , Indicadores e Reagentes , Cinética , Dados de Sequência Molecular , Especificidade por Substrato , Temperatura
19.
Biochim Biophys Acta ; 1243(3): 315-24, 1995 Apr 13.
Artigo em Inglês | MEDLINE | ID: mdl-7727505

RESUMO

The novel alkaline amylopullulanase produced by alkalophilic Bacillus sp. KSM-1378 was purified to an electrophoretically homogeneous state from culture medium. The purified enzyme was a glycoprotein with an apparent molecular mass of about 210 kDa and an isoelectric point of pH 4.8. The N-terminal amino acid sequence was Glu-Thr-Gly-Asp-Lys-Arg-Ile-Glu-Phe-Ser-Tyr-Glu-Arg-Pro and showed no homology to the N-terminal regions of other amylopullulanases reported to date. The enzyme was able to attack specifically the alpha-1,6 linkages in pullulan to generate maltotriose as the major end product, as well as the alpha-1,4 linkages in amylose, amylopectin and glycogen to generate various oligosaccharides. The pH and temperature optima for the pullulanase and alpha-amylase activities were pH 9.5 and 50 degrees C and pH 8.5 and 50 degrees C respectively. Both activities were strongly inhibited by well characterized inhibitors, such as diethyl pyrocarbonate and N-bromosuccinimide. The pullulanase activity was specifically inactivated by Hg2+ ions, alpha-cyclodextrin and beta-cyclodextrin while the amylase activity was strongly inhibited by EDTA and EGTA, although inhibition could be reversed by Ca2+ ions. It is suggested that the single alkaline amylopullulanase protein has two different active sites, one for the cleavage of alpha-1,4-linked substrates and one for the cleavage of alpha-1,6-linked substrates.


Assuntos
Bacillus/enzimologia , Proteínas de Bactérias/metabolismo , Glicosídeo Hidrolases/metabolismo , Sequência de Aminoácidos , Amilopectina/metabolismo , Amilose/metabolismo , Proteínas de Bactérias/química , Proteínas de Bactérias/isolamento & purificação , Configuração de Carboidratos , Ciclodextrinas/farmacologia , Estabilidade Enzimática , Glucanos/metabolismo , Glicogênio/metabolismo , Glicosídeo Hidrolases/química , Glicosídeo Hidrolases/isolamento & purificação , Concentração de Íons de Hidrogênio , Hidrólise , Ponto Isoelétrico , Mercúrio/farmacologia , Dados de Sequência Molecular , Peso Molecular , Fragmentos de Peptídeos/química , Especificidade por Substrato , Temperatura , Trissacarídeos/metabolismo
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