Dengue genome encodes a two component protease complex (NS2B-NS3pro) essential for the viral maturation/infectivity， thus representing a key drug target. Previously， due to its "complete insolubility"， the isolated NS3pro could not be experimentally studied and it remains elusive what structure it adopts without NS2B and why NS2B is indispensable. Here as facilitated by our previous discovery， the isolated NS3pro has been surprisingly deciphered by NMR to be the first intrinsically-disordered chymotrypsin-like fold， which exists in a loosely-packed state with non-native long-range interactions as revealed by paramagnetic relaxation enhancement (PRE). The disordered NS3pro appears to be needed for bind... More
Dengue genome encodes a two component protease complex (NS2B-NS3pro) essential for the viral maturation/infectivity， thus representing a key drug target. Previously， due to its "complete insolubility"， the isolated NS3pro could not be experimentally studied and it remains elusive what structure it adopts without NS2B and why NS2B is indispensable. Here as facilitated by our previous discovery， the isolated NS3pro has been surprisingly deciphered by NMR to be the first intrinsically-disordered chymotrypsin-like fold， which exists in a loosely-packed state with non-native long-range interactions as revealed by paramagnetic relaxation enhancement (PRE). The disordered NS3pro appears to be needed for binding a human host factor to trigger the membrane remodeling. Moreover， we have in vitro refolded the NS3pro in complex with either NS2B (48-100) or the full-length NS2B (1-130) anchored into the LMPC micelle， and the two complexes have similar activities but different dynamics. We also performed molecular dynamics (MD) simulations and the results revealed that NS2B shows the highest structural fluctuations in the complex， thus providing the dynamic basis for the observation on its conformational exchange between open and closed states. Remarkably， the NS2B cofactor plays a central role in maintaining the correlated motion network required for the catalysis as we previously decoded for the SARS 3CL protease. Indeed， a truncated NS2B (48-100;Δ77-84) with the flexible loop deleted is able to trap the NS2B-NS3pro complex in a highly dynamic and catalytically-impotent state. Taken together， our study implies potential strategies to perturb the NS2B-NS3pro interface for design of inhibitors for treating dengue infection.
