Crystal structure and binding of compounds 27 and 29 to DENV3 RdRp.

<p>(A) Overview of DENV3 RdRp bound to compound <b>29</b> (boxed), displayed as ribbons with the palm subdomain in red, finger subdomain in cyan, thumb subdomain in olive and compound <b>29</b> shown as yellow sticks. The lower left panel shows a view from “the bottom of the RdRp structure” with respect to the usual “front view”. (B) Close-up view of compound <b>29</b> binding site in the N-pocket. A difference electron density map with Fourier coefficients Fo-Fc where the compound was omitted from the phase calculation is overlaid and contoured at 3 σ level. (C) Residues lining the N-pocket for <b>29</b> are shown as cyan sticks with the compound shown as yellow sticks. Individual residues are labelled according to their numbering in the DENV3 polymerase and H-bond interactions are indicated with dashed lines. Two-dimensional ligand-interaction maps of co-crystals of DENV3 (D) and DENV2 (E) RdRp domains, bound with compounds <b>29</b> and <b>27</b>, respectively, were generated using Molecular Operating Environment. Polar residues are colored light purple, charged residues have an additional blue ring and lipophilic residues are green. The degree of solvent exposure is shown by the blue halos. H-bond interactions to the amino acid side chain or main chain are shown as dashed green or blue arrows respectively, pointing towards the H-bond acceptor. Water-mediated contacts are shown as gold dashed lines. (F) Binding of <b>27</b> to the DENV4 RdRp using surface plasmon resonance in a two-fold serial dilution from a top concentration of 2.5 μM. Raw data (coloured lines) was globally fitted to a simple 1:1 binding model (black lines). (G) Multiple alignment of amino acid residues of compound binding pocket (coloured in red and green) across different members of the Flavivirus family derived from Clustal Omega program [<a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1005737#ppat.1005737.ref042" target="_blank">42</a>]. Phylogenetic tree representing their relatedness is shown in Fig 4 in <a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1005737#ppat.1005737.s001" target="_blank">S1 Text</a>. Strictly conserved amino acids are marked with an asterisk below the sequences (and in red font), semi-conserved residues with a colon (and in green font) whilst poorly conserved residues are marked with a dot.