The Study On Structure Of RhNDPK-A Protein And Primary Investigation Of Its Immunological Effects

Objietives: To study the structure of rhNDPK-A protein and then provide the basis tomake the further study of its biological functions. To evaluate the effects of rhNDPK-Aprotein on immune system and investigate the possible mechanisms.Methods: rhNDPK-A protein was separated and purified by DEAE-anion exchange,Cibacron Blue affinity chromatography and Sephadex G-25desalting. The subunit weight,the NDP Kinase activity, the purity and the polymer weight of rhNDPK-A protein weredetermined, polymer weight of rhNDPK-A protein was examined by SE-HPLC coulpledwith OPTILABDSP. In addition, the external structure is observed by AFM. The purifiedprotein was used to immunize Australian rabbits for the preparation of antisera and thereacting tite, the specifity of the antisera were determined by westerblotting. The effects ofrhNDPK-A on the survival normal mice spleen cell and synergetic effects of it on theproliferation stipulated by mitogen including PHA, ConA and LPS. The effects ofrhNDPK-A on the cell cycle of spleen cell were also detected by FCM.Results: 1. The purified rhNDPK-A protein existes mainly in the form of hexamers. 2. Invitro, rhNDPK-A protein can directly promote the survival of spleen cell and can synergizethe proliferation stipulated by PHA and ConA. 3. rhNDPK-A protein can obviouslyinfluence the cell cycle of spleen cell. 4. The reacting tite with NDPK-A was 1:800 and theantisera was specific.Conclusions: The purified rhNDPK-A protein exists mainly in the form of hexamer;rhNDPK-A protein can promote the survival of spleen cell and may take part in theadjustment of cell growth by the way of changing cell cycle.