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Evaluated the transduction ability of a prokaryotic expression protein transduction domain of TAT fusion protein which was prepared by two different methods
Pages: 466-471,455
Year: Issue:  6
Journal: Journal of Zhenjiang Medical College

Keyword:  protein transduction domaindialysisrefoldinginclusion bodiesefficacy of transduce through the cells membrance;
Abstract: Objective:To evaluate the efficacy and subcellular localization of the fusion protein eGFP-interest protein including protein transduction domain of TAT by multi-technology and to obtain the more effective cellular uptake protein for the further research of its biological function.Methods: Induced the Expression of fusion protein,and extracted and purified it from the inclusion bodies.Jurkat cells exposure to fusion protein which was directly desalted and refolded via dialysis,respectively.Flow cytometry analysis,confocal microscope observation and Western blot detection were used to evaluate the efficacy of the fusion protein to transduce into the cell cytoplasm. Results: The results of the detected by flow cytometry,confocal microscope and Western blot indicated that the fusion protein which desalted by PD-10 column could penetrate Jurkat cells more efficiently,contrasty,the renaturated protein via dialysis had low activity. Conclusion: Both of the fusion proteins treated with two different methods could translocate through the cell membranes,but the dialyzed fusion protein had low efficiency and didn′t locate into cell nucleus,would disbennifit for the following research work about the biologic activity and behavior of the fusion protein on transcriptional level.
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