Myricetin allosterically inhibits Dengue NS2B-NS3 protease as studied by NMR and MD simulations

Dengue NS2B-NS3 protease existing in equilibrium between the active and inactive forms is essential for virus replication, thus representing a key drug target. Here Myricetin, a plant flavonoid, was characterized to non-competitively inhibit Dengue protease. Further NMR study identified the protease residues perturbed by binding to Myricetin, which were utilized to construct the Myricetin-protease complexes. Strikingly, in the active form Myricetin binds a new allosteric site (AS2) far away from the active site pocket and allosteric site (AS1) for binding Curcumin, while in the inactive form it binds both AS1 and AS2. To decipher the mechanism for the allosteric inhibition by Myricetin, we conducted molecular dynamics (MD) simulations on different forms of Dengue NS2B-NS3 protease. Unexpectedly, the binding of Myricetin to AS2 is sufficient to disrupt the active conformation by displacing the characteristic NS2B C-terminal {beta}- hairpin from the active site pocket. By contrast, the binding of Myricetin to AS1 and AS2 results in locking the inactive conformation. Therefore Myricetin represents the first small molecule which allosterically inhibits Dengue protease by both disrupting the active conformation and locking the inactive conformation. The results enforce the notion that a global allosteric network exists in Dengue NS2B-NS3 protease, which is susceptible to allosteric inhibition by small molecules such as Myricetin and Curcumin. As Myricetin has been extensively used as a food additive, it might be directly utilized to fight the Dengue infections and as a promising starting for further design of potent allosteric inhibitors. Graphic Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=117 SRC="FIGDIR/small/472523v1_ufig1.gif" ALT="Figure 1"> View larger version (29K): org.highwire.dtl.DTLVardef@ec52d1org.highwire.dtl.DTLVardef@1313dc4org.highwire.dtl.DTLVardef@1ce40b5org.highwire.dtl.DTLVardef@1fa6051_HPS_FORMAT_FIGEXP M_FIG C_FIG