High-resolution structure of eukaryotic Fibrillarin interacting with Nop56 amino-terminal domain

Analysis of cross-species complex formation between eukaryotic and archaeal Fibrillarin and Nop56/Nop5 proteins. (A) Overlay of size-exclusion chromatograms of Sc Nop1^83–327 (blue), Pf Nop5^1–123 (pink), and Pf Nop5^1–123 together with Sc Nop1^83–327 (black). Peak 1 corresponds to Sc Nop1^83–327 and peak 2 to Pf Nop5^1–123. (B) Overlay of size-exclusion chromatograms of Sc Nop56^1–166 (cyan), Pf Fibrillarin (green), and Sc Nop56^1–166 together with Pf Fibrillarin (black). (C) 4%–15% Mini-PROTEAN TGX Precast Protein Gel (BioRad) of the chromatograms shown in panels A–F. The color code above the lanes indicates the proteins present in the injection mixture: Sc Nop1^83–327 (blue), Sc Nop56^1–166 (cyan), Sc Nop56^1–166-WF (orange), Pf Fibrillarin (green), and Pf Nop5^1–123 (pink). Blue/pink lanes represent fractions collected from peak 1 and peak 2 of the black chromatogram of A. The fraction loaded in the middle lane corresponds to the region between peaks 1 and 2. The green/cyan lane represents the elution peak of the black chromatogram of Figure 3B. The blue/orange lane represents the main elution peak from the black chromatogram of Figure 3D. The green/orange lane represents the elution peak from the black chromatogram of Figure 3F. Lane M contains the protein standard. (D) Overlay of size-exclusion chromatograms of Pf Fibrillarin (green), Pf Nop5^1–123 (pink), Sc Nop1^83–327 (blue), Sc Nop56^1–166 (cyan), Pf Fibrillarin with Pf Nop5^1–123 (gray), and Sc Nop1^83–227 with Sc Nop56^1–166 (black). (E) Overlay of size-exclusion chromatograms of Sc Nop1^83–327 (blue), Sc Nop56^1–166-WF mutant (orange), and the mixture of Sc Nop1^83–327 and Sc Nop56^1–166-WF mutant (black). (F) Overlay of size-exclusion chromatograms of Pf Fibrillarin (green), Sc Nop56^1–166-WF mutant (orange), and the mixture of Pf Fibrillarin and Nop56^1–166-WF mutant (black).