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20171
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Subject: Animal Biochemistry × Author: RAJIV KISHORE, K.V. × End date: 2017 × Start date: 2017 × Thesis Type: M.V.Sc. ×
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    ThesisItemOpen Access
    ISOLATION AND PURIFICATION OF GLUTATHIONE-S-TRANSFERASE (GST) AND ITS ROLE ON OXIDATIVE STRESS IN HEPATIC TISSUES OF MAREK’S DISEASE AFFECTED CHICKEN
    (SRI VENKATESWARA VETERINARY UNIVERSITY TIRUPATI - 517 502. (A.P.) INDIA, 2017-01) RAJIV KISHORE, K.V.; PADMAJA, K(MAJOR); ESWARA PRASAD, P; PUNYA KUMARI, B
    ABSTRACT: A study was conducted to isolate and purify Glutathione-S-transferase (GST) and to determine its role on oxidative stress in liver tissues of Marek’s disease (MD) affected chicken. Ten healthy and MD suspected layer birds of 35-40 weeks age for each group were procured from poultry farms in Chittoor district of A.P. The postmortem examination of MD affected birds revealed enlarged liver with discrete grayish nodules of various sizes. Histopathological examination of liver revealed extensive proliferation and infiltration of pleomorphic cells. The purified DNA samples from liver tissue of MD suspected birds were further used for PCR analysis which was found to be positive yielding a 314 bp product indicating the presence of two copies of 132 bp tandem repeat and a 50 bp primer sequence. The liver samples positive for MD were used for further analysis. The results showed a significant increase of TBARS (179.82%) in liver of MD affected chicken compared to control. Glutathione (GSH) levels were significantly reduced (52%) in liver of MD affected chicken compared to control. A significant decrease in SOD (32%), catalase (43%), GPx (55%) and GST (51%) activities were observed in MD affected chicken compared to control. The GST enzyme was isolated, purified and characterized from liver tissue of both healthy and MD affected layer chicken using glutathione agarose affinity chromatography. The enzyme was purified to 2.38 fold with a yield of 32.27% in healthy chicken whereas in MD affected chicken it was purified to 4.54 fold with a yield of 41.98%. The analysis of active fractions using SDS-PAGE showed GST-α expression with a molecular weight of 25.6kDa in both healthy and MD affected liver. In addition GST-π was expressed in MD affected chicken with a MW of 27 kDa. These results confirm that Marek’s disease in chicken induces oxidative stress which in turn leads to alterations in antioxidant enzymes. The oxidative stress induced in liver is responsible for GST π expression as a defense mechanism in MD affected chicken.