N-terminal of ?-Synuclein Involved in Regulation of Mitochondrial Function / 中国生物工程杂志

ABSTRACT

Objective: To identify the functional domain of ?-Synuclein in affecting mitochondrial function and how the function to be impaired,especially,the mitochondrial membrane potential and the release of Cytochrome c.Methods: Harvest of ?-Syn-N and ?-Syn-△N by PCR,then subcloned into the pCMV-Myc mammalian expression vector.The recombinant plasmids were transfected into HEK293T cells by Lipofectamine 2000.After detecting the protein expression by Western blot,the functional domain was detected by co-immunoprecipitation.The mitochondrial membrane potential through flow cytometry and immunofluorescence,at the same time,the release of Cytochrome c through flow cytometry to detect.Results: The recombinant plasmids were constructed successfully.CO-IP has proved that N-terminal may be the functional domain of ?-Synuclein in affecting mitochondria.Over-expression of N-terminal could depolarize the mitochondrial membrane potential and induce the Cytochrome c releasing in MN9D cells.Conclusion: N-terminal may be the functional domain of ?-synuclein and over-expression of N-terminal could decrease mitochondrial activity.