High fidelity of yellow fever virus RNA polymerase.
Three consecutive plaque purifications of four chimeric yellow fever virus-dengue virus (ChimeriVax-DEN) vaccine candidates against dengue virus types 1 to 4 were performed. The genome of each candidate was sequenced by the consensus approach after plaque purification and additional passages in cell culture. Our data suggest that the nucleotide sequence error rate for SP6 RNA polymerase used in the in vitro transcription step to initiate virus replication was as high as 1.34 x 10(-4) per copied nucleotide and that the error rate of the yellow fever virus RNA polymerase employed by the chimeras for genome replication in infected cells was as low as 1.9 x 10(-7) to 2.3 x 10(-7). Clustering of beneficial mutations that accumulated after multiple virus passages suggests that the N-terminal part of the prM protein, a specific site in the middle of the E protein, and the NS4B protein may be essential for nucleocapsid-envelope interaction during flavivirus assembly.
Pugachev
Konstantin V
KV
Acambis, Inc., Cambridge, Massachusetts 02139, USA.
Guirakhoo
Farshad
F
Ocran
Simeon W
SW
Mitchell
Fred
F
Parsons
Megan
M
Penal
Caroline
C
Girakhoo
Soheila
S
Pougatcheva
Svetlana O
SO
Arroyo
Juan
J
Trent
Dennis W
DW
Monath
Thomas P
TP
D012321:DNA-Directed RNA Polymerases / Q000378:metabolism*; D003715:Dengue / Q000517:prevention & control; D003716:Dengue Virus / Q000235:genetics; D017422:Sequence Analysis, DNA; D012692:Serial Passage; D014158:Transcription, Genetic; D014614:Vaccines, Synthetic; D010948:Viral Plaque Assay; D014765:Viral Vaccines; D019065:Virus Assembly; D014779:Virus Replication; D015005:Yellow fever virus / Q000201:enzymology / Q000235:genetics
D014614:Vaccines, Synthetic; D014765:Viral Vaccines; D012321:DNA-Directed RNA Polymerases