Structural Studies Into The TSLP/Anti-TSLP Antibody Complex

The expression of thymic stromal lymphopoietin,TSLP,is widely distributed,including epithelial cells on the surface of the lung,skin and intestinal barrier.TSLPsare up-regulated after allergen stimulation,which promotes the maturation of DC cells(dendritic cells).Mature DC cells can induce allergic asthma.Through the study of genomics,TSLP was found to be a relevant site for allergic inflammation.In mouse allergen-induced asthma models and primate allergen-induced asthma models,TSLP has been proved to play a key role in inducing allergic asthma.In this experiment,TSLP-and WM35-Fab(TSLP antibody)-expressing transient and stable plasmids were constructed.After the confirmation of transient transfection that TSLP and WM35-Fab can be successfully expressed in 293E cells,TSLP and WM35-Fab were constructed to make stable CHO cells lines.In order to facilitate purification,a His tag was attached to the C-terminus of TSLP and WM35-Fab protein.TSLP and WM35-Fab high-purity proteins were obtained by Ni column affinity chromatography and size exclusion chromatography.The TSLP and WM35-Fab proteins are expressed in stable CHO cell lines,and the homogeneity is high.After trying different incubation ratios,it was found that the WM3 5-Fab/TSLP protein complex showed the best incubation effect when the WM35:TSLP molar ratio was 1:3.In the laboratory,44 screening kits were used to screen the crystallization conditions of W35-Fab/TSLP protein complex.Crystalsfrom three growing conditions showed good quality and were subjected to X-ray diffraction experiments by which the diffraction data were collected.In summary,in this study we obtained the crystallization conditions of W35-Fab/TSLP protein complex,and collected the crystal structure data with high resolution,which laid the foundation for further structural studies.