Human parainfluenza virus 3 (HPIV3) belongs to the
Paramyxoviridae
, causing annual worldwide epidemics of respiratory diseases, especially in newborns and infants. The core components consist of just three viral proteins: nucleoprotein (N), phosphoprotein (P), and RNA polymerase (L), playing essential roles in replication and transcription of HPIV3 as well as other paramyxoviruses. Viral genome encapsidated by N is as a template and recognized by RNA-dependent RNA polymerase complex composed of L and P. The offspring RNA also needs to assemble with N to form nucleocapsids. The N is one of the most abundant viral proteins in infected cells and chaperoned in the RNA-free form (N
0
) by P before encapsidation. In this study, we presented the structure of unassembled HPIV3 N
0
in complex with the N-terminal portion of the P, revealing the molecular details of the N
0
and the conserved N
0
-P interaction. Combined with biological experiments, we showed that the P binds to the C-terminal domain of N
0
mainly by hydrophobic interaction and maintains the unassembled conformation of N by interfering with the formation of N-RNA oligomers, which might be a target for drug development. Based on the complex structure, we developed a method to obtain the monomeric N
0
. Furthermore, we designed a P-derived fusion peptide with 10-times higher affinity, which hijacked the N and interfered with the binding of the N to RNA significantly. Finally, we proposed a model of conformational transition of N from the unassembled state to the assembled state, which helped to further understand viral replication.
IMPORTANCE
Human parainfluenza virus 3 causes annual epidemics of respiratory diseases, especially in newborns and infants. For the replication of HPIV3 and other paramyxoviruses, only three viral proteins are required: phosphoprotein (P), RNA polymerase (L), and nucleoprotein (N). Here, we reported the crystal structure of the complex of N and its chaperone P. We described in details how P acts as a chaperone to maintain the unassembled conformation of N. Our analysis indicated that the interaction between P and N is conserved and mediated by hydrophobicity, which can be used as a target for drug development. We obtained a high-affinity P-derived peptide inhibitor, specifically targeted N and greatly interfered with the binding of the N to RNA, thereby inhibiting viral encapsidation and replication. In summary, our results provide new insights into the paramyxovirus genome replication and nucleocapsid assembly, and lay the basis for drug development.
Structural Basis for the Drug Extrusion Mechanism by a MATE Multidrug Transporter and Peptide Drug Development
