Construction and secetory expression of gene engineering bacteria of uric acid enzyme

Abstract:

Objective：To clone uricase encoding gene in Candida utilis into the modified expression vector of pET-28a，and to make the uricase gene in Escherichia coli（E.coli） continuously express in secreted form with biological activity. Methods：To construct ex－pression vector pET-28a，RT-PCR method was used to delete the Lac O and Lac I sequence of pET-28a. The target gene was fused by the the N-terminal of uricase encoding gene and the signal peptide of protein A in Staphylococcus aureus. The fusion gene was cloned into the expression vector pET-28a，then transformed into E.coli DH5ɑ and expressed. The biological activity of expression product was identified and measured by SDS-PAGE gel electrophoresis. Results：The uricase gene in E.coli was continuously ex－pressed with biological activity in secreted form. Conclusion：As the normal flora in the gut，E.coli can be used to construct the ex－pression vector pET-28a-sUOX，which has secrete protein effect. Expression of uricase can be applied to the hyperuricemia model，which provide a basis for reducing uricase level through intestine.