Figure 1.
Comparison of LSD1/CoREST X-ray structure and heterogeneous conformations from conformational clustering of molecular dynamics trajectories.
Left column: X-ray structure of LSD1/CoREST bound to the H3-histone N-terminal tail (PDB ID: 2V1D); LSD1 (orange cartoons), CoREST (cyan cartoons), H3-tail (purple spheres), and the FAD cofactor (green tubes) are highlighted. LSD1/CoREST has a well-characterized amine oxidase domain (AOD) that binds the H3-histone N-terminal tail and demethylates the fourth lysine residues of the H3-histone N-terminal tail. Connected to the AOD is the SWIRM domain crucial for substrate recognition [26]. A unique feature of LSD1 is the Tower domain that serves as interface for associating with CoREST, and is required for nucleosome binding. Middle column: MD centroids of the reduced unbound conformational ensemble. Right column: MD centroids of the reduced H3-histone N-terminal tail-bound conformational ensemble. MD centroids are color coded from red (high centroid rank) to blue (low centroid rank).
Figure 2.
Druggable Site Visualizer (DSV) workflow and graphical interface.
The DSV Visualize function allows the user to easily view predicted binding sites on multiple receptor structures by mapping FTMap Consensus Sites (CSs) and SiteMap sites on a receptor and displaying the results with Visual Molecular Dynamics (VMD). DSV automated processing of an ensemble of structures displays the predicted binding sites onto a representative user-defined reference structure (in this case the top ranked MD centroid). After sourcing the DSV script, a single text on the command line instructs DSV to load the reference structure, align the remaining centroids to the reference, and then display FTMap CSs as spheres and SiteMap sites as surfaces. The spheres are sized according to CS rank and colored according to their centroid rank. The SiteMap surfaces are also colored according to centroid rank.
Figure 3.
Druggable Site Visualizer (DSV) mapping of the top FTMap consensus sites (CSs).
FTMap CSs are shown as red spheres on X-ray structures (Panel A) and as spheres colored according to representative MD centroids (Panel B). In each panel, the top row displays the top five CSs reported by FTMap; the bottom row displays the top 10 CSs. DSV displays sphere size correlated to FTMap CS rank. For graphical purposes, the FTMap CSs from all representative MD centroids are mapped onto the structure of the highest ranked MD centroid. If present during the FTMap mapping calculations, the H3-histone N-terminal tail (present in 2V1D and MD Bound) and the SNAIL1 N-terminal peptide (present in 2Y48) are highlighted as purple spheres. The H3-histone N-terminal tail and SNAIL1 N-terminal peptide were rendered more transparent when there was overlap with FTMap spheres. Solid arrows highlight new CSs at the AOD/SWIRM interface not observed in the X-ray structures and hollow arrows highlight new CSs at the AOD/Tower interface not observed in the X-ray structures (cf. 2V1D H3-Absent with MD Bound H3-Absent and 2V1D H3-Present with MD Bound H3-Present).
Figure 4.
Comparison between FTMap and SiteMap druggability mapping.
The top-five FTMap consensus sites (red spheres) and SiteMap sites (surfaces) are displayed as calculated for X-ray and MD structures. DSV Visualize was used to display the FTMap and SiteMap results, coloring the SiteMap sites according the MD centroid rank. In all cases SiteMap surfaces overlap with FTMap spheres and predict druggable regions beyond FTMap predicted space.
Figure 5.
SiteMap sites on LSD1/CoREST overlap with crystal contact regions. Panel A shows the crystal-packing interactions of three LSD1/CoREST molecules represented as red, blue, and grey opaque surfaces. In Panels B and C LSD1/CoREST are represented as transparent surfaces and SiteMap sites are colored surfaces to correspond with the unit in Panel A, e.g. the blue SiteMap sites in Panel B originate from the blue LSD1/CoREST in Panel A.
Figure 6.
Crystallographic data of Pro-Leu-Ser-Phe-Leu-Val binding to a region on the surface of LSD1/CoREST AOD.
The peptide (green) and LSD1 (brown) are highlighted (nitrogen atoms: blue; oxygen atoms: red). The unbiased 2Fo-Fc electron density map (contoured at 1.2 σ level) was calculated prior to inclusion of the peptide in the refinement. Residue Pro1 of the bound peptide was not visible in the electron density and, therefore, was not included in the model. See also Table 1 and Figure 8, region D.
Figure 7.
MD reduced ensemble shows SiteMap sites predicted small peptide binding to LSD1 AOD.
Panel A: SiteMap surfaces from H3-bound MD centroids are mapped onto the structure of LSD1/CoREST bound to Pro-Leu-Ser-Phe-Leu-Val peptide (purple spheres: PDB code 3ZMV). DSV Visualize was used to map the SiteMap sites from MD bound centroids onto 3ZMV with SiteMap surfaces colored according to centroid rank. The close-up views show SiteMap sites overlapping with the bound peptide. Panel B: Residues Arg312 and Phe320 are key for Pro-Leu-Ser-Phe-Leu-Val peptide binding and SiteMap predictions. Left: The X-ray conformation of Arg312 and Phe320 (Figure 6); SiteMap failed to predict a hot spot in this region after removal of the peptide coordinates. Middle: Instead, Arg312 and Phe320 adopted conformations that opened the Pro-Leu-Ser-Phe-Leu-Val binding pocket leading to SiteMap predictions of a clear hot spot. Right: MD snapshots in which residues Arg312 and Phe320 adopted conformations that closed binding site and prevented the prediction of SiteMap hot spots in the Pro-Leu-Ser-Phe-Leu-Val binding site.
Figure 8.
Potential small-molecule and peptide LSD1/CoREST druggable sites.
The five most favorable druggable regions thus far unexplored are highlighted using colored spheres and named based on their location: SANT2/Tower interface (green, A), AOD/Tower interface (red, B), SWIRM/AOD interface (blue, C), Peptide binding region (yellow, D), and small AOD pocket (grey, E). LSD1 (orange), CoREST (cyan), and the H3-histone N-terminal tail (purple) are shown as cartoons. Table 1 summarizes the residues in each identified druggable region. See also Figure S2 for a description of the Select-residues criterion used for analysis.
Table 1.
LSD1/CoREST residues within each predicted druggable region.