Design of NPL.
A, the chemical structure of sulfo-SBED and the partners of the three functional groups. The NHS group (green box) is for covalent linkage with nanobodies (Nb-1053, surface structure). The p-azido benzamido group (blue box) is for photosensitive linkage with the candidate interaction proteins (IPs), and biotin (red box) is for affinity purification with streptavidin beads. The distance between the succinimidyl group and azido group is around 14.3 Å. In the crystal structure of Nb-1053 (PDB code 5F1K), the CDRs are colored yellow, and the four lysines are colored red. B, protocol and schematic of the NPL approach. The CD38/Nb-1053 complex is the actual crystal structure (PDB code 5F1K) we solved previously. The four red stars represent the biotins in sulfo-SBED, which were transferred from Nb-1053 to CD38, and its interaction proteins.