Refolding of reduced/denatured ribonuclease by high performance liquid chromatography

    HUANG Yaª²Yu, TONG Wei, GUO Liª²An, GENG Xinª²Du

    1Department of Hematology, 2Department of Medical Affairs, Xijing Hospital, 3Departmant of Chemistry, School of Pharmacy, Fourth Military Medical University, Xi¬ðan 710033, China, 4Departmant of Chemistry, Northª²West University, Xi¬ðan 710069, China

    ¡¾Abstract¡¿ AIM: To investigate the refolding of the reduced/denatured ribonuclease (RNase) by high performance hydrophobic interaction chromatography (HPHIC) and high performance ion exchange chromatography (HPIEC) and to study the factors influencing its refolding. METHODS: The reduced/denatured RNase by guanidine hydrochloride or urea with high concentration in the presence of dithiotheitol (DTT) was first oxidized by oxidizedª²gluthatione (GSSG) in solution and then refolded by liquid chromatography. RESULTS: In the presence of low concentration of denaturing agent and a suitable concentration of GSSG in the mobile phrase, the bioactivity and mass recoveries of the refolded RNase increased. CONCLUSION: Compared with other elution methods, both HPHIC and HPIEC, with the advantages of high renaturation efficiency, fast process and simple operation, have good potentials in application. ......