ISOLATION AND IDENTIFICATION OF URIC ACID DEGRADING BACTERIA, OPTIMIZATION OF URICASE PRODUCTION AND PURIFICATION OF URICASE ENZYME.

Uricase plays an important role in the medical field, such as in biochemical diagnosis, where it is utilized as a reagent for uric acid detection in serum, urine and other biological fluids and in biosensors that can readily detect uric acid more specific, accurate and sensitive than the other methods. Furthermore, uricase can be used as a protein drug for reducing toxic urate accumulation in diseases such as gout. The aim of the present work is to isolate, identify and purify uricase enzyme from bacteria as well as optimize the medium and cultural conditions for maximum enzyme production. The uricase producing bacteria was isolated from different environments in and around Ernakulam Dist, Kerala. Uricase activity was initially identified by the formation of halo zone in uric acid incorporated medium. The potential uricase producing strain was then identified by cultural, morphological and biochemical analysis. Further, the identity of the strain was confirmed by PCR reaction of 16srDNA primers, sequencing and then by the BLAST analysis. The results showed that the isolated bacteria had highest homology (99%) with Ochrobactrum anthropi.The isolate was then evaluated by submerged fermentation for enzyme production. Submerged fermentation process was carried out under different pH, temperature, salinity and under different carbon and nitrogen sources to find out the optimum conditions. The optimization studies showed that the maximum enzyme activity was (0.0409 IU/mg protein) at 40 ºC temperature. The enzyme produced under all the optimum conditions was partially purified by ammonium sulphate precipitation, dialysis and DEAE ion exchange chromatography. The final purification after DEAE ion exchange chromatography yielded specific activity and fold purification of 0.0455IU/mg protein and 1.26 respectively.