Novel, structure-based mechanism for uridylylation of the genome-linked peptide (VPg) of picornaviruses.
The VPg peptide, which is found in poliovirus infected cells either covalently bound to the 5'-end of both plus and minus strand viral RNA, or in a uridylylated free form, is essential for picornavirus replication. Combining experimental structure and mutation results with molecular modeling suggests a new mechanism for VPg uridylylation, which assigns an additional function, that of scaffold, to the polymerase. The polarity of the NMR structure of VPg is complementary to the binding site on the surface of poliovirus polymerase determined previously by mutagenesis. Docking VPg at this position places the reactive tyrosinate close to the 5'-end of Poly(A)7 RNA when this is bound with its 3'-end in the active site of the polymerase. The triphosphate tail of a UTP moiety, base paired with the 5'-end of the RNA, projects back over the Tyr3-OH and is held in position by conserved positively charged side-chains of VPg. Other conserved residues mediate binding to the polymerase surface and serve as ligands for metal ion catalyzed transphosphorylation. Additional viral proteins or a second polymerase molecule may aid in stabilizing the components of the reaction. In the model complex, VPg can direct its own uridylylation before entering the polymerase active site.
Sealy Center for Structural Biology, Department of Biochemistry and Molecular Biology, University of Texas Medical Branch, Galveston, Texas 77555-0857, USA. chschein@utmb.edu
Catherine H
CH
Schein
David E
DE
Volk
Numan
N
Oezguen
Aniko
A
Paul
D000595:Amino Acid Sequence; D001483:Base Sequence; D001665:Binding Sites; D017124:Conserved Sequence; D016679:Genome, Viral; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D019906:Nuclear Magnetic Resonance, Biomolecular; D010849:Picornaviridae / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D017955:Poliovirus / Q000737:chemistry / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D012324:RNA-Dependent RNA Polymerase / Q000737:chemistry / Q000378:metabolism; D012261:Ribonucleoproteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D014529:Uridine / Q000737:chemistry / Q000378:metabolism*; D017361:Viral Nonstructural Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
D012261:Ribonucleoproteins; D017361:Viral Nonstructural Proteins; D012324:RNA-Dependent RNA Polymerase; D014529:Uridine