[Screening, characterization and expression of microbial urate oxidase]

Area of Science:

• Biochemistry
• Enzymology
• Molecular Biology

Background:

• Urate oxidase (Uox) is crucial for uric acid metabolism, with applications in diagnostics and treating hyperuricemia and gout.
• Identifying novel Uox sources with high production efficiency is essential for biotechnological advancements.

Purpose of the Study:

• To isolate and characterize a high-producing urate oxidase (Uox) from a novel bacterial strain.
• To elucidate the biochemical properties and structural features of the isolated Uox.
• To enable heterologous expression of the Uox gene for future applications.

Main Methods:

• Isolation and identification of bacterial strain OUC-1 using 16S rRNA gene sequencing.
• Purification and characterization of Uox enzyme, including optimal pH, temperature, and kinetic analysis (Km).
• Gene amplification, sequencing, 3D structure prediction (SWISS-MODEL), and heterologous expression in E. coli.

Main Results:

• Bacterial strain OUC-1, identified as Bacillus fastidiosus, was isolated as a high Uox producer.
• Purified Uox exhibited optimal activity at pH 10.0 and 40 °C, with a Km of 0.15±0.04 mmol/L for uric acid.
• Uox activity was enhanced by Mg²⁺ and inhibited by Zn²⁺ and SDS; the gene was successfully cloned and expressed in E. coli.

Conclusions:

• Bacillus fastidiosus OUC-1 is a promising source for efficient urate oxidase production.
• The characterized Uox enzyme possesses favorable properties for potential therapeutic and diagnostic uses.
• Successful heterologous expression provides a foundation for enzyme engineering and large-scale production.