Figure 3.

Comparison of the TLC structures obtained from two different crystal forms. Molecules were superimposed using the C^α positions of the 58 conserved β-­barrel residues (as in Fig. 1 ▶ b). (a) Superposition of TLC crystallized in space group P2₁ (chain A, blue) and in space group C2 (PDB code 1xki; gold, with modelled residues in grey; Breustedt et al., 2005 ▶). The bound 1,4-butanediol and the hydrogen-bonded water molecule are represented as spheres, while the conserved disulfide bond is shown in stick representation (white). Most of the β-strands are elongated towards the open end of the β-barrel in the P2₁ structure. (b) Pairwise C^α-atom distances between the two different TLC crystal structures after superposition of the 58 conserved C^α positions of the β-barrel, resulting in an overall C^α r.m.s.d. value of 1.86 Å (β-strands A–H are labelled as bars; loops 1, 2, 3 and 4 connect strands A and B, C and D, E and F, and G and H, respectively). The largest deviations between the two crystal structures occur in β-strands B, C and D and in the neighbouring loops 1 and 2 at the open end of the β-barrel. (c) The electrostatic interactions that may trigger the two alternative conformations of loop 1. In the P2₁ structure (dark grey) residues Glu27 and Lys108 (green) at the tips of the adjacent loops 1 and 4 make an electrostatic contact that fixes the ‘open’ conformation of loop 1 (blue). In contrast, the ‘closed’ conformation of this loop in the C2 structure (light grey; loop 1 coloured gold, modelled residues coloured yellow) is stabilized by a similar interaction between residues Glu34 and Lys114 (orange). (d) Flexibility of the region around the disulfide bridge. While the disulfide bond between Cys61 and Cys153 is well ordered in the P2₁ structure (dark grey; loop 2 and the C-terminal peptide segment coloured blue, side chains coloured violet), only residue Cys61 was resolved in the C2 structure (light grey; loop 2 and the C-terminal peptide segment coloured gold, modelled residues coloured yellow, Cys side chain coloured orange) and appears shifted outward by 5.6 Å compared with the P2₁ structure (red dotted line between the corresponding C^α positions). Nevertheless, the C^α distance between Cys61 in the C2 structure and Cys153 in the superimposed P2₁ structure is only 0.3 Å larger than the distance between Cys61 and Cys153 within the P2₁ structure (green dotted lines). By assuming a more extended backbone conformation at residue Glu151 (side chain displayed in blue), the (unresolved) C-terminal peptide segment of the C2 structure could easily span the 0.3 Å distance and move Cys153 sufficiently close to Cys61, indicating that the disulfide bond is probably also formed in the C2 crystal (despite the differing conformation of loop 2) but is not visible in the electron density owing to local structural disorder.