Separate molecules of West Nile virus methyltransferase can independently catalyze the N7 and 2′-O methylations of viral RNA cap

Abstract: West Nile virus methyltransferase catalyzes N7 and 2′-O methylations of the viral RNA cap (GpppA-RNA→m7GpppAm-RNA). The two methylation events are independent, as evidenced by efficient N7 methylation of GpppA-RNA→m7GpppA-RNA and GpppAm-RNA→m7GpppAm-RNA, and by the 2′-O methylation of GpppA-RNA→GpppAm-RNA and m7GpppA-RNA→m7GpppAm-RNA. However, the 2′-O methylation activity prefers substrate m7GpppA-RNA to GpppA-RNA, thereby determining the dominant methylation pathway as GpppA-RNA→m7GpppA-RNA→m7GpppAm-RNA. Mutant enzymes with different methylation defects can trans complement one another in vitro. Furthermore, sequential treatment of GpppA-RNA with distinct methyltransferase mutants generates fully methylated m7GpppAm-RNA, demonstrating that separate molecules of the enzyme can independently catalyze the two cap methylations in vitro. [Copyright &y& Elsevier]