An RNA cap (nucleoside-2'-O-)-methyltransferase in the flavivirus RNA polymerase NS5: crystal structure and functional characterization.
Viruses represent an attractive system with which to study the molecular basis of mRNA capping and its relation to the RNA transcription machinery. The RNA-dependent RNA polymerase NS5 of flaviviruses presents a characteristic motif of S-adenosyl-L-methionine-dependent methyltransferases at its N-terminus, and polymerase motifs at its C-terminus. The crystal structure of an N-terminal fragment of Dengue virus type 2 NS5 is reported at 2.4 A resolution. We show that this NS5 domain includes a typical methyltransferase core and exhibits a (nucleoside-2'-O-)-methyltransferase activity on capped RNA. The structure of a ternary complex comprising S-adenosyl-L-homocysteine and a guanosine triphosphate (GTP) analogue shows that 54 amino acids N-terminal to the core provide a novel GTP-binding site that selects guanine using a previously unreported mechanism. Binding studies using GTP- and RNA cap-analogues, as well as the spatial arrangement of the methyltransferase active site relative to the GTP-binding site, suggest that the latter is a specific cap-binding site. As RNA capping is an essential viral function, these results provide a structural basis for the rational design of drugs against the emerging flaviviruses.
Egloff
Marie-Pierre
MP
Architecture et Fonction des Macromolécules Biologiques, UMR 6098 CNRS et Université Aix-Marseille I et II, ESIL, Campus de Luminy, F-13288 Marseille Cedex 09, France.
Benarroch
Delphine
D
Selisko
Barbara
B
Romette
Jean-Louis
JL
Canard
Bruno
B
D000595:Amino Acid Sequence; D001665:Binding Sites; D018360:Crystallography, X-Ray; D004305:Dose-Response Relationship, Drug; D006160:Guanosine Triphosphate / Q000378:metabolism; D008780:Methyltransferases / Q000378:metabolism*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009711:Nucleotides / Q000737:chemistry; D011485:Protein Binding; D017510:Protein Folding; D017434:Protein Structure, Tertiary; D012313:RNA / Q000378:metabolism*; D013997:Time Factors; D017361:Viral Nonstructural Proteins / Q000737:chemistry* / Q000502:physiology*
C096041:NS5 protein, flavivirus; D009711:Nucleotides; D017361:Viral Nonstructural Proteins; D012313:RNA; D006160:Guanosine Triphosphate; D008780:Methyltransferases; C032117:cap I RNA (nucleoside-2'-)methyltransferase; C032116:cap II RNA(nucleoside-2'-)methyltransferase