hpp10sod融合蛋白表达及其抗氧化功效分析-expression of hpp 10od fusion protein and its antioxidant activity analysis.docx

AbstractObjective：Superoxide dismutase, a kind of protease, which can clear oxygen free radical, and it is the key molecules of defensing the toxicity of oxygen free radical. Studies have shown that SOD has a series of action, such as anti-aging, immunity adjustment, resisting radiation damage, and skin care and beauty. It can be used as medicine enzyme, but also be used as cosmetics and health products. But because of its big molecular weight, the absorption rate remains to be improved by oral or spread on skin. Cell membranepenetration peptide can act as a new carrier with Cu·Zn-SOD into the cytoplasm ornucleus, improving the absorption efficiency of SOD.Methods: (1) Construction of recombinant plasmid of pET15b-SOD, pET15b-hPP10– SOD;（2）Obtain the purified protein of SOD and hPP10–SOD. We identify the correctness by Westernbloting. (3) We identify the original efficiency of the penetrating the cell membrane by immunofluorescence, and the accurate efficiency by Westernbloting. (4)The SOD enzyme of hPP10–SOD activity detection is detected by the kids itself. (5)We detect the cell activity by MTT-tset, when the cell was dealed with hPP10–SOD.Results: (1) We get the recombinant plasmid of pET15b-SOD, pET15b-hPP10–SOD;We obtain purified fusion protein of SOD and hPP10–SOD, and the protein is correct.The immunofluorescence test results show that the hPP10–SOD can penetrat the cell membrane obviously when the concentration is 5 μM, the Westernblot test results show that the effect of penetrating the cell membrane is resed along with the concentration increasing within a certain range, and it can keep 24 hours. (4) It has a very good antioxidant activity when the concentration of hPP10-SOD is 10μM (p 0.05). (5) The MTT-tset results that the impact on cell activity has been small when the concentration of hPP10-SOD is reached 10μM.Conclusion: hPP10, new original cell membrane penetrating peptides, is effective penetrating the cell membrane. The results indicate that the