Purificação e caracterização parcial de uma B-D glicosidade de Artemia franciscana com ação sobre celobiose e lactose

-D-glucosidase (EC 3.2.1.21) is one of the most interesting glycosidases, especially for hydrolysis cellobiose releasing glucose, is last step degradation of cellulose. This function makes the -D-glucosidase is of great interest as a versatile industrial biocatalyst, being critical to various bio-treatment / biorefinery processes, such as bioethanol production. Hen in the report, a -D-glucosidase was extracts from protein extracted of the invertebrate marine Artemia franciscana was purified and characterized with a combination of precipitation with ammonium sulfate (0 - 30%, 30 to 50%, 50 to 80%), the fraction saturated in the range of 30 to 50% (called F-II) was applied in a molecular exclusion chromatography, in Sephacryl S-200, the fractions corresponding to the first peak of activity of -D-glucosidase were gathered and applied in a chromatography of ion exchange in Mono Q; the third peak this protein obtained chromatography, which coincides with the peak of activity of -D-glucosidase was held and applied in a gel filtration chromatography Superose 12 where the first peak protein, which has activity of -D-glucosidase was rechromatography on Superose 12. This enzyme is probably multimerica, consisting of three subunit molecular mass of 52.7 kDa (determined by SDS-PAGE) with native molecular mass of 157 kDa (determined by gel filtration chromatography on Superose 12 under the system FPLC). The enzyme was purified 44.09 times with a recovery of 1.01%. Using up p-nitrophenyl-β-D-glucopiranoside as substrate obtained a Km apparent of 0.229 mM and a Vmax of 1.109 mM.60min-1.mL-1mM. The optimum pH and optimum temperature of catalysis of the synthetic substrate were 5.0 and 45 °C, respectively. The activity of the -D-glucosidase was strongly, inhibited by silver nitrate and N- etylmaleimide, this inhibition indicates the involvement of radical sulfidrila the hydrolysis of synthetic substrate. The -D-glucosidase of Artemia franciscana presented degradativa action on celobiose, lactose and on the synthetic substrate -nitrophenyl-β-D-glucopiranoside indicating potential use of this enzyme in the industry mainly for the production of bioethanol (production of alcohol from the participating cellulose), and production hydrolysate milk (devoid of milk lactose)

Purificação e caracterização parcial de uma B-D glicosidase de Artemia franciscana com ação sobre celobiose e lactose

-D-glucosidase (EC 3.2.1.21) is one of the most interesting glycosidases, especially for hydrolysis cellobiose releasing glucose, is last step degradation of cellulose. This function makes the -D-glucosidase is of great interest as a versatile industrial biocatalyst, being critical to various bio-treatment / biorefinery processes, such as bioethanol production. Hen in the report, a -D-glucosidase was extracts from protein extracted of the invertebrate marine Artemia franciscana was purified and characterized with a combination of precipitation with ammonium sulfate (0 - 30%, 30 to 50%, 50 to 80%), the fraction saturated in the range of 30 to 50% (called F-II) was applied in a molecular exclusion chromatography, in Sephacryl S-200, the fractions corresponding to the first peak of activity of -D-glucosidase were gathered and applied in a chromatography of ion exchange in Mono Q; the third peak this protein obtained chromatography, which coincides with the peak of activity of -D-glucosidase was held and applied in a gel filtration chromatography Superose 12 where the first peak protein, which has activity of -D-glucosidase was rechromatography on Superose 12. This enzyme is probably multimerica, consisting of three subunit molecular mass of 52.7 kDa (determined by SDS-PAGE) with native molecular mass of 157 kDa (determined by gel filtration chromatography on Superose 12 under the system FPLC). The enzyme was purified 44.09 times with a recovery of 1.01%. Using up p-nitrophenyl-β-D-glucopiranoside as substrate obtained a Km apparent of 0.229 mM and a Vmax of 1.109 mM.60min-1.mL-1mM. The optimum pH and optimum temperature of catalysis of the synthetic substrate were 5.0 and 45 °C, respectively. The activity of the -D-glucosidase was strongly, inhibited by silver nitrate and N- etylmaleimide, this inhibition indicates the involvement of radical sulfidrila the hydrolysis of synthetic substrate. The -D-glucosidase of Artemia franciscana presented degradativa action on celobiose, lactose and on the synthetic substrate -nitrophenyl-β-D-glucopiranoside indicating potential use of this enzyme in the industry mainly for the production of bioethanol (production of alcohol from the participating cellulose), and production hydrolysate milk (devoid of milk lactose)

Estudo da imunoexpressão dos transportadores de glicose 1 e 3 em carcinomas epidermóides de lábio inferior e sua relação com parâmetros clínico-patológicos

The Epidermoid Carcinoma (EC) is the most common lesions located in the region of the head and neck and, despite advances in treatment modalities, the prognosis is still poor. The malignant cells show an increase in glucose uptake, process mediated by glucose transporters (GLUTs). Increased expression of GLUT 1 and GLUT 3 is related to the aggressive behavior of this lesion. The aim of this study was to evaluate, through immunohistochemistry, the expression of GLUTs 1 and 3 in EC of the lower lip. The sample consisted of 40 cases of EC of the lower lip, of which 20 had regional lymph node metastasis and the remaining 20 with absence of metastasis. The percentages of immunostained cells in front of tumor invasion and in the center of tumor were evaluated. These results were related to the presence and absence of lymph node metastasis, TNM classification and histological grading. The percentage of cytoplasmic/membranous expression of GLUT 1 ranged from 77.35% to 100%, while for GLUT 3 this value ranged from 0.79% to 100%. As for nuclear staining for GLUT 1, this percentage ranged from 0 to 0.42%, however. GLUT 3 showed only one case with nuclear staining. Despite the significant expression of tumor cells related to the proteins studied, we observed no statistically significant relationship between the variables and the antibodies analyzed, regardless of the region evaluated. However, there was a moderate positive correlation between cytoplasmic/membranous immunoexpressions of GLUT 1 in invasion front and in the tumor center (r = 0.679, p <0.001). Similarly, moderate positive correlation was found between the nuclear immunoexpressions of GLUT 1 in the invasion front and in the tumor center (r = 0.547, p <0.001). For GLUT 3, was also observed a moderate statistically significant positive correlation between cytoplasmic/membranous expression in tumor invasion front and in tumor center (r = 0.589, p <0.001). We also observed that the immunoreactivity for GLUT 1 was higher than GLUT 3 expression in invasion front (p <0.001) and tumor center (p <0.001). From these results, this study suggests that tumor hypoxia is a remarkable characteristic of the EC of the lower lip and GLUT 1 may be primarily responsible for glucose uptake into the interior of the malignant cells