Fig. 4: Structure of the complex of Fap2-ECD and hTIGIT-ECD.

a Cryo-EM density map of Fap2-ECD/hTIGIT-ECD at 6.0 Å resolution. b Comparison of the cryo-EM densities of the membrane-distal tip regions of Fap2-ECD/hTIGIT-ECD (left) and Fap2-ECD (right), as indicated by the box in (a). The Fap2 model is shown in orange for both maps. c Docking of hTIGIT-ECD (green) to Fap2-ECD (orange) with HADDOCK. The best docking pose is shown and is in agreement with the cryo-EM density. d Proposed interaction site of Fap2 and hTIGIT as indicated by the box in (c). The loop G66-P67-G68 of hTIGIT intercalates into a gap in Fap2, and main chain and side chain interactions as identified in docking and verified by MD simulations are highlighted. e Fluorescence micrographs that show HEK293 cells transfected with hTIGIT-eGFP and binding of Fap2-expressing E. coli (WT and mutant K368P_S371A_R427A_W540A). Red: E. coli labeled with CellBrite Fix 555, green: hTIGIT-eGFP. Scale bar: 50 µm. The images are representatives of data sets of 75 micrographs each. f Quantification of experiments in (e), with average integrated fluorescence signal of E. coli bound to HEK293T transfected with hTIGIT-eGFP determined from 75 micrographs each. The integrated fluorescence of E. coli was detected with the abundance of hTIGIT-eGFP signal as a prerequisite. The mean is shown as red dot and the boxes represent the interquartile range, with the median shown as horizontal line within the box. The distribution of data points is visually represented by violin plots. Statistical evaluation was done using the t-test from the ggsignif¹⁰⁵ package and visualized with ggstatplot⁸² in R.