Study On Optimization Of Coupled Procedure Of Superoxide Dismutase Purification And Modification

Conventional protein chemical modification is a complex,long period and the relative random modify process, easily lead to protein denaturation. This subject proposed a coupled procedure of protein purification and modification. The coupled procedure can simplify the original production steps, shorten the operation time, reduce the loss of protein activity, lower investment costs and production costs.This subject chose Q Sepharose Fast Flow as solid-phase medium, chose superoxide dismutase as protein model, chose monomethoxypolye- thylene glycol 5000 as modifier. This subject investigated the influencing factors on adsorption capacity, investigated the influencing factors on modification, investigated the influencing factors on elution, investigated the stability of the modified superoxide dismutase.This subject investigated adsorption factors of superoxide dismutase in chapter II. The optimum conditions of adsorption were found as followsing: the adsorption time was 30min, pH was 9.0, the concentration of superoxide dismutase was 3.0mg/ml, the temperature was room temperature. The maximum adsorption capacity for superoxide dismutase could reach 22.5mg/ml.This subject investigated the influencing factors on elution in the coupled procedure of superoxide dismutase purification and modification in chapter III. The optimum conditions of modification were found as followsing: the modification time was 270 min, the concentration of borate buffer solution was 40mmol/L, the molar ratio of monomethoxypolyethylene glycol 5000 to superoxide dismutase was 50:1, the temperature was room temperature. Two optimum ways of elution: one is gradient elution of NaCl concentration(0–1.0mol/L), another is eluted by 0.15mol/L NaCl and gradient elution of NaCl concentration (0.15–0.6mol/L).This subject investigated the thermal stability, pH stability, anti-pepsin hydrolysis and anti-trypsin hydrolysis of modifed superoxide dismutase. The results show that modified superoxide dismutase have better in temperature, acid, alkali and proteolytic enzymes.