RESUMO
RESUMEN Los residuos líquidos producidos al elaborar tinciones biológicas contienen mezclas de compuestos químicos y microorganismos, que generan un elevado impacto ambiental si no son tratados adecuadamente. Por esta razón, en el presente trabajo se evaluaron a Pleurotus ostreatus, Trametes versicolor, Enterobacter xianfangensis, Pseudomonas azotoformans, Pseudomonas sp., Bacillus subtilis y Pseudomonas fluorescens, para el tratamiento de un residuo líquido que contenía colorantes trifenilmetánicos y azóicos, a escala de laboratorio. Inicialmente, se seleccionaron las cepas con menor efecto antagónico y se determinó su potencial para producir las enzimas Lacasa, Manganeso Peroxidasa y Lignino Peroxidasa, al emplear sustratos inductores y mezclas de colorantes. Para el consorcio fúngico/bacteriano la disminución de las unidades de color y demanda química de oxígeno fueron del 99 % y 70 % a las 96 h. La remoción de estos parámetros se relacionó con la interacción positiva e incremento de las poblaciones de hongos, bacterias y la producción de enzimas ligninolíticas, obteniendo valores a las 96 h de 7.0 y 14.0 unidades logarítmicas para hongos y bacterias, con unas actividades enzimáticas de 75 U/L, 205 U/L y 0.63 U/L para Lacasa, MnP y LiP, respectivamente. Con el presente trabajo se demostró que con el uso consorcios fúngicos/bacterianos se incrementa la remoción de colorantes y se disminuye el tiempo de proceso. Sugiriendo que estos microorganismos podrían ser evaluados en plantas de tratamiento que integren diferentes unidades de tratamiento para optimizar la remoción de contaminantes con baja biodegradabilidad.
ABSTRACT The liquid wastes generated when biological stains are prepared, contain a mixture of chemical compounds and microorganisms, with high environmental impact. For this reason, Pleurotus ostreatus, Trametes versicolor, Enterobacter xianfangensis, Pseudomonas azotoformans, Pseudomonas sp., Bacillus subtilis and Pseudomonas fluorescens, were used to evaluate the treatment of a liquid waste containing triphenylmethane and azo dyes, on a laboratory scale. Initially, the strains with less antagonistic effect among them were selected for their potential to produce enzymes as Laccase, Manganese Peroxidase and Lignin Peroxidase. The enzymatic activity was determined by using inducing substrates and dye mixtures. For fungal/bacterial consortium, the decrease in color, Chemical Oxygen Demand and in Biochemical Oxygen demand was of 99 %, 70 % and 65 % at 96 h, respectively. The removal of these parameters was related to the positive interaction between the populations of fungi, bacteria and the production of ligninolytic enzymes, obtaining values of 7.0 and 14.0 logarithmic units for fungi and total bacteria at 96 h with enzymatic activities of 75 U/ L, 205 U/L and 0.63 U/L for Laccase, MnP and LiP. The present work demonstrates that using of fungal/bacterial consortia, the removal of dyes is increased, and the process time is decreased. Suggesting that these microorganisms could be evaluated in treatment plants that integrate different treatment units to optimize the removal of contaminants with low biodegradability.
RESUMO
The sulfatase family involves a group of enzymes with a large degree of similarity. Until now, sixteen human sulfatases have been identified, most of them found in lysosomes. Human deficiency of sulfatases generates various genetic disorders characterized by abnormal accumulation of sulfated intermediate compounds. Mucopolysaccharidosis type II is characterized by the deficiency of iduronate 2-sulfate sulfatase (IDS), causing the lysosomal accumulation of heparan and dermatan sulfates. Currently, there are several cases of genetic diseases treated with enzyme replacement therapy, which have generated a great interest in the development of systems for recombinant protein expression. In this work we expressed the human recombinant IDS-Like enzyme (hrIDS-Like) in Escherichia coli DH5α. The enzyme concentration revealed by ELISA varied from 78.13 to 94.35 ng/ml and the specific activity varied from 34.20 to 25.97 nmol/h/mg. Western blotting done after affinity chromatography purification showed a single band of approximately 40 kDa, which was recognized by an IgY polyclonal antibody that was developed against the specific peptide of the native protein. Our 100 ml-shake-flask assays allowed us to improve the enzyme activity seven fold, compared to the E. coli JM109/pUC13-hrIDS-Like system. Additionally, the results obtained in the present study were equal to those obtained with the Pichia pastoris GS1115/pPIC-9-hrIDS-Like system (3 L bioreactor scale). The system used in this work (E. coli DH5α/pGEX-3X-hrIDS-Like) emerges as a strategy for improving protein expression and purification, aimed at recombinant protein chemical characterization, future laboratory assays for enzyme replacement therapy, and as new evidence of active putative sulfatase production in E. coli.
