Study On Crystal Structure And Function Of The V Domain Of Human Nectin-2

Nectins are immunoglobulin-like cell adhesion molecules (CAMs) which participate in Ca2+- independent intracellular interactions. The nectin family consist of four members, which are nectin-1, -2, -3, -4 respectively. Members of nectins share similar properties in the extracellular fragment which posess three successive Ig-like domains (V, C2, and C2) in the extracellular fragment. Cell adhension are mediated through cis-dimerization and tans-dimerization between nectins. As a member of nectin family, in addition to the property and function mentioned above, nectin-2 (CD112) also plays roles in mediating virus into cells and immune defense against abnormal cells after binding with its receptor DNAM-1 (CD226), an activating receptor on NK cells and cytotoxic lymphocyte cells(CTL).To date, crystal structures of two nectin like molecules, Necl-1 and Necl-5, have been published. In view of its great importance in cell-cell adhesion and fighting against tumor cells, the crystal structure of nectin-2 will help us to understand the mechanism in cell adhesion and find the exact sites for binding with proteins expressed on virus surface and its receptor CD226.The V domain of nectin-2 was expressed in E. coli as inclusion bodies, which were subsequently refolded and purified. Lastly we got the soluble proteins and concentrated it to 5mg/ml and 10mg/ml. The soluble protein was crystallized using the hanging-drop vapor-diffusion method. The crystals diffracted to 1.85 ? resolution and belonged to the space group P21, with unit cell parameters a = 52.3 ?, b = 43.9 ?, c = 56.1 ?, andβ= 118.2°Here we report crystal structure of the v domain of nectin-2 and it was the first structure of nectin family. The protein crystallized as a tans-dimmer. Compared with the V domain of other two Necl molecule, a distinct extended loop can be easily found, which may be the key site for nectin-2 to bind with it receptor CD226. We will prepare the tetramer of the V domain of nectin-2 and use it to bind with CD226 expressed on COS7 cells to demonstrate our hypothesis. Some related experiment is undergoing.