RESUMO
Background: Catalase (CAT) is an important enzyme that degrades H2O2 into H2O and O2. To obtain an efficient catalase, in this study, a new strain of high catalase-producing Serratia marcescens, named FZSF01, was screened and its catalase was purified and characterized. Results: After optimization of fermentation conditions, the yield of catalase produced by this strain was as high as 51,468 U/ml. This catalase was further purified using two steps: DEAE-fast flow and Sephedex-G150. The purified catalase showed a specific activity of 197,575 U/mg with a molecular mass of 58 kDa. This catalase exhibited high activity at 2070°C and pH 5.011.0. Km of the catalase was approximately 68 mM, and Vmax was 1886.8 mol/min mg. This catalase was further identified by LCMS/MS, and the encoding gene was cloned and expressed in Escherichia coli BL21 (DE3) with a production of 17,267 ± 2037 U/ml. Conclusions: To our knowledge, these results represent one of the highest fermentation levels reported among current catalase-producing strains. This FZSF01 catalase may be suitable for several industrial applications that comprise exposure to alkaline conditions and under a wide range of temperatures.
Assuntos
Serratia marcescens/enzimologia , Catalase/metabolismo , Recombinação Genética , Serratia marcescens/genética , RNA Ribossômico 16S , Cinética , Catalase/isolamento & purificação , Catalase/genética , Cromatografia Líquida , Análise de Sequência de DNA , Eletroforese , Escherichia coli/genética , Escherichia coli/metabolismo , Fermentação , Peróxido de Hidrogênio/metabolismoRESUMO
A field experiment established in 1980 was conducted to evaluate the effects of open drainage ditch applied for water removal on bacterial and fungal communities of cold waterlogged paddy soils in 2011. In this experiment, traditional plate counting and temperature gradient gel electrophoresis were employed to characterize the abundance and diversity of soil bacterial and fungal communities. Four different distances from the open drainage ditch, 5, 15, 25 and 75 m with different degrees of drainage were designed for this study. Maximum populations of culturable aerobic bacteria and fungi were at 15-m distance while minimum populations were at 75-m distance. Significant differences (p < 0.05) in fungal populations were observed at all distances from open drainage ditch. The highest diversity of the bacterial community was found at a distance of 25 m, while that of the fungal community was observed at a distance of 5 m. Sequencing of excised TGGE bands indicated that the dominant bacteria at 75-m distance belonged to anaerobic or microaerobic bacteria. Relationships between microbial characteristics and soil physicochemical properties indicated that soil pH and available nitrogen contents were key factors controlling the abundance of culturable aerobic bacteria and fungi, while soil water capacity also affected the diversity of fungal community. These findings can provide the references for better design and advanced management of the drainage ditches in cold waterlogged paddy soils.
Assuntos
Biota , Bactérias/classificação , Bactérias/isolamento & purificação , Fenômenos Químicos , Fungos/classificação , Fungos/isolamento & purificação , Microbiologia do Solo , Análise por Conglomerados , Temperatura Baixa , Eletroforese em Gel de Gradiente Desnaturante , DNA Bacteriano/química , DNA Bacteriano/genética , DNA Fúngico/química , DNA Fúngico/genética , DNA Ribossômico/química , DNA Ribossômico/genética , Drenagem , Genes de RNAr , Concentração de Íons de Hidrogênio , Dados de Sequência Molecular , Nitrogênio/análise , Filogenia , RNA Bacteriano/genética , RNA Fúngico/genética , /genética , /genética , Análise de Sequência de DNA , Homologia de Sequência do Ácido Nucleico , Solo/químicaRESUMO
Eight silver-staining protocols were applied to detect DNA in polyester-backed gels to select the optimal. Results showed important differences in staining quality and that four methods were well-suited for TGGE gels due to high sensitivity and low background, including the Bassam et al. methods, the manufacturer method and our improved method.