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. 2020 Oct 1;295(50):17027–17045. doi: 10.1074/jbc.RA120.014625

Figure 3.

Figure 3.

Varied structures and ligand occupancy between chains in the FUT8:GDP complex. The four chains in the FUT8:GDP asymmetric unit A, exhibited varied GDP occupancy between the respective chains and different loop conformations (chains A, B, C, and D in the crystal lattice are labeled and boxed regions represent the GDP-binding sites shown in panel B). Chains A and B show full occupancy of the bound GDP, whereas chain C shows partial occupancy, and chain D shows no occupancy as indicated by the difference density map (Fo − Fc) for the donor analog contoured at 3σ. B, zoom-in representations of the GDP-binding sites. Upper panels, zoom-in of the difference density maps (Fo − Fc) for the donor analog as indicated by the boxed regions in panel A with cartoon representations of the respective protein regions. Middle panels, chains A-D and the apo-FUT8 structure (PDB 2DE0 (36)) are shown in cartoon representation (chain A, tan; chain B, green; chain C, slate; chain D, yellow; Apo-FUT8 2DE0, orange). Significant differences in conformation were observed in three loops (Loop 1 (residues 245-273, green), Loop 2 (residues 365-378, red), and Loop 3 (residues 436-443, cyan)) with labels in the respective color. In chains C and D, disorder in Loops 2 and 3 were observed and the ends of the respective ordered regions are indicated by the red (Loop 2) or cyan (Loop 3) spheres, respectively. The 2DE0 structure also had a disordered region for Loop 2 indicated by the red spheres. GDP (yellow sticks) is modeled in the binding site of each chain for reference. Lower panels, surface representation of the four chains in FUT8:GDP and the apo-FUT8 (2DE0) structures with GDP (yellow sticks) modeled in the binding site of each chain for reference. Coloring is as shown in the middle panels. C, the progressive conformational changes in Loops 1-3 are illustrated by an overlay of the loops from chains A-D and the PDB 2DE0 structure. A fully extended “flipped-out” conformation is represented by the 2DE0 structure and progressive closure of Loop 2 and Loop 3 and repositioning of Loop 1 in chains C and D leads to partial occupancy in the GDP-binding site for chain C. Further closure of Loops 1-3 in chains A and B leads to full occupancy of the donor analog and completion of the induced fit donor interactions.