Figure 5.

Lack of association between$\mathbf{PR}{\mathbf{D}}_{\mathbf{703}–\mathbf{800}}^{\mathbf{Strep}}$and globular ALIX domains.¹H_N/¹⁵N chemical shift perturbation profiles of (A) nonphosphorylated and (B) phosphorylated ²H/¹⁵N-labeled Bro1 on addition of three-molar equivalent of nonlabeled $\mathrm{P}\mathrm{R}{\mathrm{D}}_{\mathrm{703}–\mathrm{800}}^{\mathrm{S}\mathrm{t}\mathrm{r}\mathrm{e}\mathrm{p}}$. C, ¹H_N/¹⁵N chemical shift perturbation profiles of ¹⁵N-labeled $\mathrm{P}\mathrm{R}{\mathrm{D}}_{\mathrm{703}–\mathrm{800}}^{\mathrm{S}\mathrm{t}\mathrm{r}\mathrm{e}\mathrm{p}}$ on addition of nonlabeled Bro1 (blue), phosphorylated Bro1 (red), V (black), and Bro-V (dark green); one at a time, $\mathrm{P}\mathrm{R}{\mathrm{D}}_{\mathrm{703}–\mathrm{800}}^{\mathrm{S}\mathrm{t}\mathrm{r}\mathrm{e}\mathrm{p}}$ to globular ALIX domains molar ratio = 1:3. Region highlighted in semitransparent orange rectangle indicates the location of the C-terminal strep purification tag (residues 801–808). ALIX, apoptosis-linked gene-2 interacting protein X; PRD, proline-rich domain.