Figure 1. The crystal structure of the C-terminal region of HUWE1 reveals an asymmetric dimer.

(A) Structural organization of HUWE1 (3951–4374), shown for molecule A. The bi-lobal catalytic HECT domain is colored yellow, the previously uncharacterized region orange. (B) Superposition of the HECT domains of molecules A and B (grey). The N-terminal regions are highlighted. (C) Asymmetric dimer formed of molecules A and B in the crystal. The C-terminal residues (4370–4374, ‘C-tail’) of molecule B are highlighted in magenta. Two loop regions in molecule B, distant from the dimer interface, are disordered (dotted lines) (top). Cartoon model of the asymmetric dimer (bottom). The C-lobe and the C-tail (magenta) of molecule B are in a locked conformation (see Figure 2C,D). In all structural figures, the protein backbones are rendered as cartoons and the side chains of relevant residues as ball-and-stick models.

DOI: http://dx.doi.org/10.7554/eLife.21036.003

Figure 1—figure supplement 1. Crystallographic B-factor analysis.

(A) Isotropic C_α B-factors are plotted onto the crystal structure of the HUWE1 dimer. The table lists the mean isotropic B-factors per molecule and region, as calculated by Phenix (Adams et al., 2010). The dimerization regions of molecules A and B show B-factors that are below average overall. For molecule B that has higher B-factors than molecule A, the dimerization region has B-factors below average; for molecule A, the opposite is true. Importantly, however, both dimerization regions are well defined in the electron density. Residues at the N-terminus of the pointer helices that belong to the expression tag, including a disconnected stretch of residues, whose chain identity could not be assigned (see Figure 3—figure supplement 2) are characterized by high B-factors. While the B-factors for molecule B are higher than for molecule A, they fall into an acceptable range of the Wilson B factor for this dataset, as validated by Phenix (Adams et al., 2010). (B) Lattice environment of the asymmetric HUWE1 dimer. Surrounding symmetry mates are shown in ribbon representation (grey). The difference in the B-factors for the two molecules may originate from the different environments of two molecules in the context of the crystal lattice. A significant portion of molecule B – the one that is characterized by relatively high B-factors – faces a large water channel. In contrast, molecule A is more tightly embedded into lattice contacts and, consistently, displays lower B-factors.

DOI: http://dx.doi.org/10.7554/eLife.21036.004