Preparation of isocitrate lyase versus etablishment of a drug screening model for isocitrate lyase inhibitors

Abstract

Objective: To prepare isocitrate lyase and establish a high throughput screening model for isocitrate lyase inhibitors. Methods: We used four-step chromatography or HiTrap Chelating HP to purify isocitrate lyase that expresses by BL21 (DE3). With isocitrate as substrate, the purified isocitrate lyase was albe to converts isocitrate to succinate and glyoxylate. Glyoxylate reacted to phenylhydrazine turn to phenylhydrazone. We could measure the light absorption value of enzyme system at 324nm to determine the isocitrate lyase activity. The screening model used to measure the activity of protein isocitrate lyase. Results: The drug screening model was established and optimized. The signal to noise ratio (S/N) of the screening model was much higher than 3, and the coefficient of variation (CV) is much smaller than 10%. Conclusion: We peparated isocitrate lyase by HiTrap Chelating HP and established a drug screening model for isocitrate lyase inhibitors with high specificity and stability. This new drug screening model may be efficiently used to screen inhibitors of isocitrate lyase.