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![[Figure 3]](ba5290fig3mag.jpg)
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Figure 3
`Hidden' alternative conformations in natural and artificial protein structures. (a) Natural protein: residues Asn173 and Arg464 in a 0.88 Å resolution structure of catalase (PDB entry 1gwe; Murshudov et al., 2002  ) are each modeled with a single conformation. However, 2Fo − Fc (0.7σ as a light blue volume and blue mesh) and ±Fo − Fc (+3.5σ and −3.5σ in green and red, respectively; both volume and mesh) electron-density maps suggest that the existing Arg464 conformation is overmodeled and reveal evidence for a `hidden' alternative conformation. Supporting this interpretation, the existing Arg464 conformation sterically clashes (red/orange/yellow spikes; Word, Lovell, LaBean et al., 1999) with several waters (red spheres) that were mistakenly modeled into that electron density. (b) A refitted and rerefined model, with the Asn173 side-chain amide flipped 180° (curved dotted arrow; Word, Lovell, Richardson et al., 1999), an alternative rotamer added for Arg464 (purple versus orange), the offending waters removed and alternative water positions that are mutually exclusive with the original Arg464 conformation added, results in a better fit to the electron density, including diminished Fo − Fc difference peaks, elimination of steric clashes and a more extensive hydrogen-bonding network (green dotted lines). Some additional partial-occupancy waters may also be present, given the remaining positive Fo − Fc density. (c) Artificial protein: residues Arg104 and Gln105 in chain B of a 2.09 Å resolution structure of a de novo designed protein (PDB entry 5e6g; Jacobs et al., 2016) are modeled with single conformations. However, 2Fo − Fc (0.7σ as a light blue volume and blue mesh) and ± Fo − Fc electron-density maps (+3.0σ and −3.0σ in green and red, respectively; both volume and mesh) reveal evidence for a `hidden' alternative conformation for Arg104 and a missing partial-occupancy ordered water molecule nearby that were not specified in the design model (arrows). (d) A refitted and rerefined model, with alternative conformations (purple versus orange) for Arg104 and the partial-occupancy water added, results in a better fit to the electron density, including diminished Fo − Fc difference peaks and a more extensive hydrogen-bonding network (green dotted lines). Images were obtained using PyMOL (Schrödinger). |
![[Figure 3]](ba5290fig3.jpg)
 | STRUCTURAL BIOLOGY |
ISSN: 2059-7983
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