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Figure 5
FoFo difference maps reveal local conformational shifts connecting the active site, interdomain inter­face, and dimer inter­face. Center: overview of the isomorphous FoFo difference electron-density map at ±3σ (green–red mesh) for the 240 K data set (cyan) minus the 100 K data set (dark blue) (see Fig. S10 for FoFo maps for all tem­per­atures). Ligands from cocrystal structures are shown at the active site (dashed oval) (pale orange, 6lu7), interdomain inter­face (purple, 5ree; yellow, 5rec), and dimer inter­face (orange, 7lfp; pink, 5rf0). (a) Glu290 switches from one side-chain rotamer at 100 K to two alternate rotamers at 240 K. Glu290 is spatially adjacent to Cys128, which switches from two alternate rotamers at 100 and 240 K to a single rotamer at 277 K and above in our multiconformer models; the alternate rotamer occupancy is lower at 240 K, consistent with its positive FoFo peak (see Fig. S12). These residues are near two ligands from separate crystallographic screens (7lfp and 5rf0), as well as many ordered PEG mol­ecules from the crystallization cocktails of various structures (7kvr, 7kvl, 7kfi, and 7lfe). (b) An ∼45°-rotated view relative to part (a) shows that these two ligands bind at the dimer inter­face of the biological monomer, constituted in the crystal from a symmetry-related protomer (gray surface). This inter­face also includes the Asp197 region (right). (c) Thr198 switches from two alternate side-chain rotamers at 100 K to a single rotamer at 240 K, while Glu240 – located across the interdomain inter­face – changes side-chain rotamer (curved arrows), with additional effects on the adjacent backbone of Pro241. In the other direction from Asp197 (down in this view), other residues in the P5 substrate-binding pocket loop (Fig. 3[link]) undergo conformational adjustments en route to the active site. Meanwhile, an inter­acting water mol­ecule at 100 K (blue sphere) becomes displaced at 240 K, and is correspondingly absent in that model.

Volume 9| Part 5| September 2022| Pages 682-694
ISSN: 2052-2525