"Resurrected" human-source urate oxidase with high uricolytic activity and stability

Area of Science:

• Biochemistry
• Evolutionary Biology
• Enzyme Engineering

Background:

• Urate oxidase (UOX) is a pseudogene in humans due to evolutionary mutations.
• Despite being a pseudogene, human UOX can be transcribed and translated.
• Higher homology to deduced human urate oxidase (dHU) may correlate with lower immunogenicity.

Purpose of the Study:

• To construct a "resurrected" human-source urate oxidase (rHU19) based on dHU.
• To evaluate the uricolytic activity and catalytic efficiency of rHU19.
• To assess the theoretical suitability of rHU19 for treating hyperuricemia and gout.

Main Methods:

• Bioinformatic analysis to deduce human urate oxidase (dHU).
• Gene resurrection and protein engineering to construct rHU19.
• Enzymatic assays to measure uricolytic activity and catalytic efficiency.

Main Results:

• rHU19 demonstrated significantly higher uricolytic activity (8.29 U/mg) and catalytic efficiency (3.32 s⁻¹ μM⁻¹) than wild porcine urate oxidase (wPU) and porcine-baboon chimera (PBC).
• rHU19 maintained high homology (93.75%) with dHU.
• The engineered enzyme showed promising characteristics for therapeutic applications.

Conclusions:

• The engineered human urate oxidase (rHU19) exhibits superior enzymatic properties compared to existing urate oxidase therapies.
• rHU19's high homology to dHU suggests potentially lower immunogenicity, making it a theoretically suitable candidate for treating gout and hyperuricemia.