Sars Coronavirus S Glycoprotein Expression In Hela Cells

Objective: To observe the subcellular orientation of spike glycoprotein of SARS-Coronovirus (S glycoprotein) while it is transfected by non-virus vector. To illustrate the possible roles that S glycoprotein plays during SARS-CoV infection. Methods: To analyze the mutation and the physical-chemical character of the S glycoprotein by the bioinformatcs, the plasmid with the S glycoprotein of specimen has been sequenced. Expression plasmids containing the full length cDNA of S glycoprotein with GFP or HA peptide sequence were constructed and transfected into HeLa cell by PolyFect Transfection Reagent. The fusion protein was detected under confocal microscope and by Western blotting. Results: The mutation progress of the S glycoprotein, of which the molecular weight is 139KD and the PI is 5.54, is very fast. Fusion protein was located evenly in transfected cells, both nucleus and cytoplast, but not segregated in cell surface. This orientation did not changed with time course. Western blotting detected a specific band at 70~90 KD. Conclusion: S glycoprotein was located in nucleus and cytoplasm but not to be assembled into cell membrane while it was transfected in HeLa cell by non-virus vector. It was not located as full length in HeLa cell. Therefore, the protease cut-site in the S glycoprotein can not be excluded.