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Expression of Related Cholesterol Dehydrogenation Protein and Bioactivity Research

ZHANG Jing;ZHANG Wen-qiang;QIN Hui-min;MAO Shu-hong;XUE Jia-lu;LU Fu-ping;Key Laboratory of Industrial Fermentation Microbiology,National Engineering Laboratory for Industrial Enzymes (NELIE) ,College of Biotechnology,Tianjin University of Science & Technology;Henan Li Wei Medicine Co. ,Ltd;  
The neverland from the Drosophila melanogaster play essential roles in cholesterol dehydrogenation. To clone the neverland from the Drosophila melanogaster c DNA library and construct the eukaryote expression plasmid p IEx-6-nvd and p XY212-nvd to expressing the recombinant NVD in S2 cell and S.cerevisiae W303-1A for further analyzing its bioactivity. Results of Western blot indicates the recombinant protein expressed by S2 cell and S. cerevisiae W303-1A can specifically combine with 6 × His antibody. Results of HPLC shows the cholestrol can be converted to 7-dehydrocholesterol by the recombinant S2 cell,however,the purified NVD and the recombinant S. cerevisiae W303-1A can not conversion cholesterol to 7-dehydrocholesterol,even the mixed recombinant S. cerevisiae W303-1A cell lysate and S2 cell lysate,neither. As the above result shows,the NVD need some chaperonins to complete the electron-transfer reactions when mediates the cholesterol dehydrogenation reaction.
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