The structural basis for cap binding by influenza virus polymerase subunit PB2.
Influenza virus mRNAs are synthesized by the trimeric viral polymerase using short capped primers obtained by a 'cap-snatching' mechanism. The polymerase PB2 subunit binds the 5' cap of host pre-mRNAs, which are cleaved after 10-13 nucleotides by the PB1 subunit. Using a library-screening method, we identified an independently folded domain of PB2 that has specific cap binding activity. The X-ray structure of the domain with bound cap analog m(7)GTP at 2.3-A resolution reveals a previously unknown fold and a mode of ligand binding that is similar to, but distinct from, other cap binding proteins. Binding and functional studies with point mutants confirm that the identified site is essential for cap binding in vitro and cap-dependent transcription in vivo by the trimeric polymerase complex. These findings clarify the nature of the cap binding site in PB2 and will allow efficient structure-based design of new anti-influenza compounds inhibiting viral transcription.
Guilligay
Delphine
D
Grenoble Outstation, European Molecular Biology Laboratory, 6 rue Jules Horowitz, BP181, 38042 Grenoble Cedex 9, France.
Tarendeau
Franck
F
Resa-Infante
Patricia
P
Coloma
Rocío
R
Crepin
Thibaut
T
Sehr
Peter
P
Lewis
Joe
J
Ruigrok
Rob W H
RW
Ortin
Juan
J
Hart
Darren J
DJ
Cusack
Stephen
S
D002460:Cell Line; D006801:Humans; D009980:Influenza A virus / Q000737:chemistry / Q000378:metabolism*; D017354:Point Mutation; D017434:Protein Structure, Tertiary; D012314:RNA Cap Analogs / Q000378:metabolism; D012315:RNA Caps / Q000378:metabolism*; D012324:RNA-Dependent RNA Polymerase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D014158:Transcription, Genetic; D014764:Viral Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*
C057027:PB2 protein, Influenzavirus A; D012314:RNA Cap Analogs; D012315:RNA Caps; D014764:Viral Proteins; C017516:7-methylguanosine triphosphate; D012324:RNA-Dependent RNA Polymerase