Molecular and proteomic identification of Arthrobacter gandavensis isolated from cows with subclinical mastitis in a dairy farm

Aims@#The genus Arthrobacter is a pleomorphic and heterogeneous Gram-positive bacteria mainly isolated from the soil, only two species of Arthrobacter have been reported worldwide as pathogens of veterinary importance. This paper aims to report the isolation and identification of the Arthrobacter gandavensis from cows with subclinical mastitis at a dairy farm in the savanna of Bogotá, Colombia.@*Methodology and results@#A total of 209 milk and skin samples were taken from cows with and without subclinical mastitis, nasal swabs from workers and the environment. All samples were cultured in blood and MacConkey agar and identified by 16S rRNA gene sequencing and mass spectrometry MALDI TOF-MS. From the isolates identified, 33 corresponded to Staphylococcus spp., nine to the Enterobacteriaceae family and seven from Arthrobacter spp. (only identified by MALDI-ToF MS). The A. gandavensis isolates were obtained from six different positive cows for the California mastitis test, all with a matching pattern corresponding to Arthrobacter gandavensis strain DSM N: 15046, isolated from milk from cows with subclinical mastitis in Belgium. Analysis of the 16S rRNA gene showed 100% genetic similarity with sequences of A. gandavensis previously reported in the NCBI databases.@*Conclusion, significance and impact of study@#The identification by MALDI-ToF-MS and molecular, as shown in this report, is important to provide data that allow us to approach the actual ecology of the opportunistic pathogens of subclinical mastitis, especially in regions where the infection is endemic.

La rizobacteria promotora del crecimiento vegetal Arthrobacter agilis UMCV2 coloniza endofíticamente a Medicago truncatula / The plant growth-promoting rhizobacterium Arthrobacter agilis UMCV2 endophytically colonizes Medicago truncatula

Arthrobacter agilis UMCV2 es una bacteria rizosférica que promueve el crecimiento vegetal de plantas leguminosas proveyéndoles hierro soluble. Un segundo mecanismo de promoción se da a través de la producción de compuestos volátiles que estimulan los mecanismos de absorción de hierro. Adicionalmente, A. agilis UMCV2 tiene la capacidad de inhibir el crecimiento de organismos fitopatógenos. En el presente trabajo se emplea una combinación de las técnicas de reacción en cadena de la polimerasa cuantitativa e hibridación in situ con fluorescencia para detectar y cuantificar la presencia de la bacteria en los tejidos internos de la planta leguminosa Medicago truncatula. Nuestros resultados demuestran que A. agilis UMCV2 se comporta como una bacteria endófita de M. truncatula especialmente en medios donde el hierro está disponible.

Arthrobacter agilis UMCV2 is a rhizosphere bacterium that promotes legume growth by solubilization of iron, which is supplied to the plant. A second growth promotion mechanism produces volatile compounds that stimulate iron uptake activities. Additionally, A. agilis UMCV2 is capable of inhibiting the growth of phytopathogens. A combination of quantitative polymerase chain reaction and fluorescence in situ hybridization techniques were used here to detect and quantify the presence of the bacterium in the internal tissues of the legume Medicago truncatula. Our results demonstrate that A. agilis UMCV2 behaves as an endophytic bacterium of M. truncatula, particularly in environments where iron is available.

Antimicrobial profile of Arthrobacter kerguelensis VL-RK_09 isolated from Mango orchards

Abstract An actinobacterial strain VL-RK_09 having potential antimicrobial activities was isolated from a mango orchard in Krishna District, Andhra Pradesh (India) and was identified as Arthrobacter kerguelensis. The strain A. kerguelensis VL-RK_09 exhibited a broad spectrum of in vitro antimicrobial activity against bacteria and fungi. Production of bioactive metabolites by the strain was the highest in modified yeast extract malt extract dextrose broth, as compared to other media tested. Lactose (1%) and peptone (0.5%) were found to be the most suitable carbon and nitrogen sources, respectively, for the optimum production of the bioactive metabolites. The maximum production of the bioactive metabolites was detected in the culture medium with an initial pH of 7, in which the strain was incubated for five days at 30 °C under shaking conditions. Screening of secondary metabolites obtained from the culture broth led to the isolation of a compound active against a wide variety of Gram-positive and negative bacteria and fungi. The structure of the first active fraction was elucidated using Fourier transform infrared spectroscopy, electrospray ionization mass spectrometry, 1H and 13C nuclear magnetic resonance spectroscopy. The compound was identified as S,S-dipropyl carbonodithioate. This study is the first report of the occurrence of this compound in the genus Arthrobacter.

Microorganisms in Vacuum Stored Flower Bee Pollen

Contamination with sanitary microorganisms from Enterobacteriaceae, Pseudomonadaceae, Staphylococcaceae, Micrococcaceae and Bacillaceae families in flower bee pollen from Bulgaria after one-year vacuum-packed cold storage has been found. Dried flower bee pollens intended for human consumption were with high incidence rate of contamination with Pantoea sp. (P. agglomerans and P. agglomerans bgp6) (100%), Citrobacter freundii (47%), Proteus mirabilis (31.6%), Serratia odorifera (15.8%) and Proteus vulgaris (5.3%). Bee pollens were also positive for the culture of microorganisms from Staphylococcaceae, Micrococcaceae and Bacillaceae families: Staphylococcus hominis subsp hominis, Staphylococcus epidermidis, Arthrobacter globiformis, Bacillus pumilis, Bacillus subtilis and Bacillus amyloliquefaciens. It was concluded that, if consumed directly, the vacuum-packed cold stored dried bee pollen, harvested according hygienic requirements from bee hives in industrial pollution-free areas without intensive crop production, is not problem for healthy human.

La bacteria Arthrobacter agilis UMCV2 y diversas aminas inhiben el crecimiento in vitro de hongos destructores de madera / Bacterium Arthrobacter agilis UMCV2 and diverse amines inhibit in vitro growth of wood-decay fungi

El reino Fungi está representado por innumerable cantidad de organismos entre los cuales se encuentran hongos patógenos que deterioran los principales componentes estructurales de la madera, como celulosa, hemicelulosa y lignina. El objetivo de nuestro trabajo fue caracterizar la actividad antifúngica y la producción de diversas aminas de Arthrobacter agilis UMCV2 con acción antagónica sobre hongos xilófagos. Para ello, se aislaron 4 organismos fúngicos (designados en conjunto UMTM) a partir de madera en descomposición en un bosque de pino encino de la comunidad de Cuanajo, Michoacán, México. Dos de ellos presentaron una clara actividad enzimática de celulasas, xilanasas y enzimas accesorias óxido-reductoras, y fueron identificados como pertenecientes a 2 géneros agresivos para la madera: Hypocrea (aislado UMTM3) y Fusarium (aislado UMTM13). In vitro, las aminas evaluadas mostraron tener efecto inhibitorio sobre el crecimiento de los UMTM y la dimetilhexadecilamina; uno de estos compuestos mostró un fuerte potencial para ser utilizado como tratamiento preventivo contra el ataque de hongos destructores de madera.

The kingdom Fungi is represented by a large number of organisms, including pathogens that deteriorate the main structural components of wood, such as cellulose, hemicellulose and lignin. The aim of our work was to characterize the antifungal activity in Arthrobacter agilis UMCV2 and diverse amines against wood-decaying fungi. Four fungal organisms (designated as UMTM) were isolated from decaying wood samples obtained from a forest in Cuanajo-Michoacán, México. Two of them showed a clear enzymatic activity of cellulases, xylanases and oxido-reducing enzymes and were identified as Hypocrea (UMTM3 isolate) and Fusarium (UMTM13 isolate). In vitro, the amines showed inhibitory effect against UMTM growth and one of the amines, dimethylhexadecylamine (DMA16), exhibited strong potential as wood preventive treatment, against the attack of decaying fungi.

Tutorial for writing systematic reviews for the Brazilian Journal of Physical Therapy (BJPT) / Tutorial para elaboração de revisões sistemáticas para o Brazilian Journal of Physical Therapy (BJPT)

Systematic reviews aim to summarize all evidence using very rigorous methods in order to address a specific research question with less bias as possible. Systematic reviews are widely used in the field of physical therapy, however not all reviews have good quality. This tutorial aims to guide authors of the Brazilian Journal of Physical Therapy on how systematic reviews should be conducted and reported in order to be accepted for publication. It is expected that this tutorial will help authors of systematic reviews as well as journal editors and reviewers on how to conduct, report, critically appraise and interpret this type of study design. .

Revisões sistemáticas têm como objetivo sumarizar toda a evidência disponível, através de métodos rigorosos, para responder a uma pergunta de pesquisa específica com o mínimo de viés possível. Revisões sistemáticas são amplamente utilizadas na fisioterapia, porém nem todas as revisões possuem boa qualidade. Esse tutorial tem como objetivo guiar os autores do Brazilian Journal of Physical Therapy sobre como revisões sistemáticas deveriam ser conduzidas e descritas para que sejam aceitas para publicação. Espera-se que esse tutorial irá auxiliar autores de revisões sistemáticas, assim como editores e revisores de periódicos em como conduzir, descrever, fazer análise crítica e interpretar esse tipo de delineamento de pesquisa.

Hexavalent chromium reduction by aerobic heterotrophic bacteria indigenous to chromite mine overburden

Microbiological analysis of overburden samples collected from chromite mining areas of Orissa, India revealed that they are rich in microbial density as well as diversity and dominated by Gramnegative (58%) bacteria. The phenotypically distinguishable bacterial isolates (130) showed wide degree of tolerance to chromium (2-8 mM) when tested in peptone yeast extract glucose agar medium. Isolates (92) tolerating 2 mM chromium exhibited different degrees of Cr+6 reducing activity in chemically defined Vogel Bonner (VB) broth and complex KSC medium. Three potent isolates, two belonging to Arthrobacter spp. and one to Pseudomonas sp. were able to reduce more than 50 and 80% of 2 mM chromium in defined and complex media respectively. Along with Cr+6 (MIC 8.6-17.8 mM), the isolates showed tolerance to Ni+2, Fe+3, Cu+2 and Co+2 but were extremely sensitive to Hg+2 followed by Cd+2, Mn+2 and Zn+2. In addition, they were resistant to antibiotics like penicillin, methicillin, ampicillin, neomycin and polymyxin B. During growth under shake-flask conditions, Arthrobacter SUK 1201 and SUK 1205 showed 100% reduction of 2 mM Cr+6 in KSC medium with simultaneous formation of insoluble precipitates of chromium salts. Both the isolates were also equally capable of completely reducing the Cr+6 present in mine seepage when grown in mine seepage supplemented with VB concentrate

Development of Arthrobacter woluwensis Bacteremia in a Patient with Multiple Myeloma: A Case Report and Comprehensive Literature Review / 감염과화학요법

Arthrobacter spp., which are coryneform gram-positive bacilli, are widely distributed in the environment, including soil. In humans, infection with Arthrobacter is recognized as an opportunistic infection. In particular, since the first reported case in 1996, human infection by A. woluwensis has been reported only four times. We report on a case of A. woluwensis bacteremia in a 76-year-old female patient with multiple myeloma. Performance of 16S rRNA gene sequence analyses resulted in identification of A. woluwensis. The patient was treated with teicoplanin, and the central venous port was removed. Since then, no growth has been observed on repeated blood cultures. The patient was discharged well after the fever subsided.

Optimization of physical parameters for lipase production from Arthrobacter sp. BGCC#490.

The physical parameters for the production of thermostable, alkaline lipase from Arthrobacter sp. BGCC# 490 were optimized using response surface methodology (RSM), employing face centered central composite design (FCCCD). The design was employed by selecting pH, temperature and incubation period as the model factors and to achieve maximum yield, interaction of these factors was studied by RSM. A second-order quadratic model and response surface method showed that the optimum conditions for lipase production (pH 10.0, temperature 40oC and incubation period 48 h) resulted in 1.6-fold increase in lipase production (13.75 EUml-1), as compared to the initial level (8.6 EUml-1) after 48 h of incubation, whereas its value predicted by the quadratic model was 12.8 EUml-1. Lipase showed stability in the pH range 8-10 and temperature range 40-60oC, with maximum activity at pH 9.0 and temperature 50oC. Lipase activity was enhanced in the presence of K+, Ca2+ and Mg2+ ions, but inhibited by Hg2+ ions. The enzyme exhibited high activity in the presence of acetone, isopropanol and ethanol, but was unaffected by methanol. These properties suggest that the lipase may find potential applications in the detergent industry. The present work also demonstrated the feasibility of using experimental design tools to optimize physical parameters for lipase production by an indigenous Arthrobacter sp.

Biodehydrogenation of 11beta-hydroxyl melroxyprogesterone by Arthrobacter simplex UR016 in microemulsion system / 生物工程学报

To improve mass transfer and enhance the yield for C(1,2) biodehydrogenation of steroid 11beta-hydroxyl medroxyprogesterone, we carried out the dehydrogenation reaction of 11beta-hydroxyl medroxyprogesterone in an oil-in-water (O/W) microemulsion by Arthrobacter simplex UR016. We studied the effects of system composition, dehydrogenation temperature and substrate concentration on microbial transformation. We formulated a suitable O/W microemulsion system with Arthrobacter simplex UR016 culture broth as aqueous phase, 10 g/L of edible oil as oil phase, 4 g/L of Tween-O80 and 7% (V/V) alcohol as surfactant and cosurfactant. The optimal dehydrogenation temperature was 33 degrees C. The results showed that in Tween-80/alcohol/edible oil/water microemulsion system, the hydrophobic steroid was solubilised and diffused effectively, with the maximum conversion rate of 88.6% at 46 h under 4 g/L substrate concentration, an increase of 66.2% compared to that in aqueous system. The C(1,2) biodehydrogenation of 11beta-hydroxyl medroxyprogesterone is more efficient in water-edible oil microemulsion system than in aqueous system.

Novel s-triazine-degrading bacteria isolated from agricultural soils of central Chile for herbicide bioremediation

s-Triazine-degrading bacterial strains were isolated from long-term simazine-treated agricultural soils of central Chile. The number of culturable heterotrophic bacteria of these agricultural soils (7 x 10(6) CFU/g of dry soil) was not affected by simazine application on field. The simazine-degrading bacterial strains P51, P52 and C53 were isolated by enrichment in minimal medium using simazine as the sole nitrogen source. Resting cells of strains P51 and P52 degraded >80 percent of simazine within 48 hrs, whereas strain C53 was able to remove >60 percent of the herbicide. The atzA and atzD genes of the s-triazine upper and lower catabolic pathways were detected in strains P51 and C53, while only atzD gene was observed in strain P52. To compare the bacterial 16S rRNA gene sequence structure, ARDRA were performed using the restriction enzymes Msp1 and Hha1. ARDRA indicated that strain P52 was a different ribotype than C53 and P51 strains. For further characterization the novel isolates were identified by 16S rRNA gene sequencing. Strains C53 and P51 belong to the genus Stenotrophomonas and the strain P52 belongs to the genus Arthrobacter . s -Triazine-degrading bacterial strains isolated from contaminated soils could be used as biocatalysts for bioremediation of these herbicides.

Cambios en la viabilidad de dos bacterias marinas antárticas expuestas a la radiación solar en la columna de agua: influencia de la mezcla vertical / Changes in viability of two Antarctic marine bacteria exposed to solar radiation in the water column: influence of vertical mixing

Se estudió el efecto de la radiación ultravioleta (RUV) sobre dos cepas bacterianas marinas antárticas (UVps y UVvi) en la columna de agua de la caleta Potter (Shetland del Sur, Antártida). Frascos de cuarzo con las cepas en estudio fueron expuestos a la radiación solar en superficie, a 1 m y a 3 m de profundidad. Se realizaron ensayos con exposición directa y con filtros interferenciales que discriminaron la radiación UVA y la UVB. En otros ensayos se simuló una mezcla vertical de 4 m/h. Ambas cepas mostraron una disminución significativa del número de unidades formadoras de colonias, tanto en superficie como a 1 m de profundidad, luego de exponerlas a dosis superficiales de UVB de 8,4 kJ m-2. El estudio con filtros interferenciales mostró una disminución significativa de la viabilidad en ambos tratamientos UV en superficie y a 1 m. La cepa UVps mostró mayor sensibilidad a la UVB que a la UVA. La mezcla vertical amortiguó el daño causado por la UVB cuando la dosis en superficie fue de 4,8 kJ m-2. Este efecto amortiguador no se observó cuando la dosis en superficie fue de 7,7 kJ m-2. Estos resultados muestran que el efecto negativo de la RUV sobre el bacterioplancton sería particularmente importante en el primer metro de profundidad de las aguas costeras antárticas con abundante material particulado en suspensión.

The effect of UV radiation on two Antarctic marine bacterial strains (UVps and UVvi) was studied in the water column of Potter Cove (South Shetland, Antarctica). Quartz flasks were filled with the bacterial suspensions and exposed to solar radiation at 0 m, 1 m and 3 m depth. Assays using flasks exposed to direct solar radiation and others using flasks covered with/by interferential filters which discriminate between UVA and UVB, were performed. In other assays, a vertical mixing of 4 m/h was simulated. Both strains showed a significant decrease in viability (expressed as colony - forming units) when exposed to a surface UVB dose of 8.4 kJ m-2. Studies with interferential filters showed a significant decrease at 0 and 1 m depth under both UV treatments. The UVps strain appeared to be more sensitive to UVB than to UVA. Damage produced by UVB was attenuated by the vertical mixing when the surface UVB dose was 4.8 kJ m-2. This effect was not observed when surface UVB dose was 7.7 kJ m-2. These results show that the negative effect caused by UVB radiation on the bacterioplankton would be significant only in the first meter of water column of the Antarctic coastal waters with high levels of suspended particulate material.

Study on the atrazine-degrading genes in Arthrobacter sp. AG1 / 生物工程学报

Atrazine could be used as the sole carbon, nitrogen and energy sources for growth by strain Arthrobacter sp. AG1, and the atrazine-degrading genes of AG1 were found to be the combination of trzN, atzB and atzC. The atrazine chloride hydrolysase gene trzN was cloned by PCR amplification,whose sequence shared 99% identity with that of Norcardioides sp. C190. Two large plasmids were found in AG1,and trzN and atzB were confirmed to be localized on the larger plasmid pAG1 by the method of southern hybridization. Subculture of AG1 in liquid LB for three generations, 34% of the subsequent cells were found to lose degrading activity, however, neither plasmid was lost. PCR amplification results showed that the mutants had only lost the trzN gene instead of atzB and atzC. It was deduced that mutation might be due to the trzN gene deletion from the plasmid. This study provided new evidence that atrazine metabolic genotypes were resulted from horizontal gene transfer between different bacteria under environmental selective pressure.

A Case of Bacteremia Caused by Arthrobacter woluwensis / 대한임상미생물학회지

Arthrobacter spp. are coryneform bacteria known as a soil flora and also part of normal flora of human. Since coryneform bacteria are often reported to be a cause of life-threatening diseases, and especially the human infections of Arthrobacter spp. are reported recently, it is important to identify them to the genus and species levels by additional studies including molecular tests. We report a case of bacteremia caused by Arthrobacter woluwensis, which was misidentified initially as Leifsonia aquatica by commercial kits and conventional tests, but correctly identified by 16S rRNA sequencing.

Uso tradicional y actual de spirulina sp. (arthrospira sp.) / Traditional and present use of spirulina sp. (arthrospira sp.)

Spirulina sp. (Arthrospira sp.) es una cianobacteria filamentosa no diferenciada, habitante de lagos alcalinos, que se cultiva para consumo humano debido a su contenido nutricional. En México, el consumo de esta cianobacteria se remonta a tiempos prehispánicos, cuando era conocida como tecuitlatl, siendo conocida como dihé por las tribus nativas de la región del lago de Chad, en Africa. En los últimos años se le han atribuido diversos efectos positivos en el tratamiento de algunos tipos de alergias, anemia, cáncer, enfermedades virales y cardiovasculares. Muchas de sus propiedades son consecuencia de la presencia de pigmentos como las ficobiliproteínas y los carotenoides, así como de otros compuestos como polisacáridos, ácidos grasos (destacando el ácido gama linoleico), proteínas, vitaminas y minerales. Las propiedades y aplicaciones de este organismo hacen de él un alimento promotor de la salud o nutracéutico

Optimization of polysaccharides production by bacteria isolated from soil

Six polysaccharide-producing bacteria, isolated from soil samples and identified as Pseudomonas and Arthrobacter (Strains 3B, 4B, 7B, 21B, 18E and 21D), were tested for the yield of polysaccharides produced during growth in two culture media: one containing glucose and the other sucrose (1, 2, 3, 4 and 5 percent). The yield was quantified measuring the viscosity of the broth, using the Poiscuille equation. The effect of temperature and pH of the culture media was investigated. The largest polysaccharide yield was obtained when the concentration of the carbon source was lower than 2 percent. Glucose and sucrose stimulated the polysaccharide production in a similar way. When the initial pH of the fermentation broth was increased from 5.0 up to 7.0, there was an increase in polysaccharide production. However, higher values (pH=8.0) caused a decrease in polysaccharide production. With regard to temperature, 30°C was shown to be optimum, since higher or lower temperatures had a negative effect on saccharide production.

Seis bactérias produtoras de polissacarídeos isoladas de amostras do solo (3B, 4B, 7B, 21B, 18E e 21D) pertencentes aos gêneros Pseudomonas e Arthrobacter, foram testadas quanto ao rendimento da produção de polissacarídeos em dois caldos de cultura: um com glicose e outro com sacarose, nas concentrações de 1, 2, 3, 4 e 5 por cento. O rendimento foi calculado medindo-se a viscosidade dos caldos, através da equação de Poiseuille. Também foi verificado o efeito da temperatura de incubação (25, 30 e 35°C) e o do pH (5, 6, 7 e 8) dos meios de cultura. Os resultados obtidos mostraram que concentrações da fonte de carbono até 2 por cento apresentaram os maiores rendimentos do polissacarídeo. Glicose e sacarose estimularam a produção de polissacarídeos de forma similar. Quando o pH inicial do caldo de fermentação aumentou de 5,0 para 7,0 ocorreu um aumento da produção do polissacarídeo. Entretanto, valores mais altos (pH=8,0) causaram uma diminuição na formação do produto. Com relação à temperatura, 30°C foi considerada ótima, pois temperaturas maiores ou menores do que esta exerceram um efeito negativo na produção do polissacarídeo.

Contenido microbiológico cultivable del tracto intestinal y polen almacenado de Apis mellifera (Hymenoptera: Apidae) / Cultured Microbiological Content of the Intestinal Tract and Stored Pollen of Apis mellifera (Hymenoptera: Apidae)

Se caracterizaron los microorganismos cultivables asociados con Apis mellifera. Las muestras fueron tomadas a partir de polen almacenado (joven y maduro) y transportado en corbículas y tracto digestivo de las abejas (forrajeras y recién nacidas). Se aislaron bacterias pertenecientes a los géneros Pseudomonas, Streptococcus, Micrococcus, Lactobacillus, Klebsiella, Proteus, Yersinia y Arthrobacter y hongos de los géneros Rhizopus, Alternaria y Epicoccum. De acuerdo a sus propiedades bioquímicas, algunas de estas bacterias pueden estar involucradas en la degradación de los compuestos de la capa externa del polen y son adquiridas por las abejas a través del alimento y contacto con otros individuos de la colmena. La presencia de los hongos se explica por su amplia distribución en el ambiente, ya que los tres géneros se encuentran comúnmente en el suelo y en las plantas que las abejas pueden seleccionar como fuente de alimento.

A Case of Catheter-Related Bacteremia by Arthrobacter woluwensis / 대한진단검사의학회지

Arthrobacter woluwensis, a catalase-positive coryneform bacterium recognized as an opportunistic pathogen, was repeatedly isolated from the blood of a 56-year-old male patient with metastatic colon cancer. The isolate was identified by various phenotypic tests and by sequencing analysis of 16S rRNA. Antimicrobial susceptibility testing was performed by E-test; the MICs to vancomcyin, cefotamine, and penicillin were 1.5 microgram/mL, >64 microgram/mL, and 4 microgram/mL, respectively. The patient was treated with vancomycin, and the subclavian catheter, which was presumed to be the source of the infection, was removed. Thereafter, repeated blood cultures did not grow the organism. The infections of human caused by A. woluwensis have not been reported previously in Korea, probably because of the difficulty of identifying Arthrobacter strains by conventional biochemical tests.

Purification and characteristics of creatininase from Arthrobacter sp / 生物工程学报

A creatininase produced from a Arthrobacter sp. was purified 145-fold by a series of steps including heat treatment, ammonium sulfate precipitation, DEAE-Cellulose ion-exchange and hydrophobic chromatography. The specific activity of the pure enzyme was 209u/mg. The subunit molecular mass of creatininase was estimated to be 33 700D by SDS-PAGE. The creatininase was stable in the pH range between 6.0 - 9.0 and below 60 degrees C . Its Km value for creatinine was estimated to be 21.14 mmol/L. The enzyme was markedly inactivated by incubation with 1 mmol/L of Hg2+, Ag2+, Li+, Cu2+ and 20 mmol/L of 1, 11-Phananthroline respectively. Activation was observed when the enzyme was incubated with 1 mmol/L of Co2+ and Mn2+.

Cloning and expression of L-N-carbamoylase gene from Arthrobacter BT801 in Escherichia coli / 生物工程学报

Hydantoin-utility-enzyme is widely used in enzymic production of various amino acids. One of its component, carbamoylase, is responsible for the conversion of N-carbamylamino acids to corresponding amino acids, which is crucial for the stereoselectivity and rate limiting. To improve the production of the enzyme, an L-N-carbamoylase gene from Arthrobacter BT801, a hydantoinase producting strain being able to convert 5-benzylhydantoin to phenylalanine, was cloned into E. coli. The gene was highly expressed in E. coli M15 under control of T5 promoter. A protein band about 44kD was detected by SDS-PAGE in the recombinant cell lysate. The objective product, which is principally in soluble form, represented 40% of total cell protein. The N-carbamoylase specific activity of the recombinant M15/pQE60- hyuC is 53 times higher than that of Arthrobacter BT801. The total biotransformation activity increased 8.1 times when. M15/pQE60-hyuC was added into the Arthrobacter BT801 reaction system. The successful expression of the enzyme is significant for the application of the hydantoinase producing strain or the enzyme thereof.