RNA dimerization plays a role in ribosomal frameshifting of the SARS coronavirus

Ishimaru, Daniella; Plant, Ewan P.; Sims, Amy C.; Yount, Boyd L.; Roth, Braden M.; Eldho, Nadukkudy V.; Pérez-Alvarado, Gabriela C.; Armbruster, David W.; Baric, Ralph S.; Dinman, Jonathan D.; Taylor, Deborah R.; Hennig, Mirko

All NMR spectra were recorded at 288 K, 295 K or 298 K on Bruker Avance 900, 800, 700 or 600 MHz spectrometer equipped with either a triple resonance inverse detection cryoprobe (800 and 700) or standard triple resonance inverse detection-probeheads. NMR experiments were performed on samples of 500 ml volume containing 0.2-1.2 mM SARS-CoV pk and stem-loop S3L2 variant RNA. Data were processed using NMRPipe (20) and analysed using Sparky (21) . One-dimensional imino proton spectra were acquired using a jump-return echo sequence (22) . Imino resonances were assigned sequence specifically from water flip-back, WATERGATE 2D nuclear Overhauser effect spectroscopy (NOESY) spectra (t mix = 150-200 ms) (23) . Typically, for the 1 H, 1 H-NOESY spectra, 384 complex points were recorded with an acquisition time of 24 ms for 1 H (o 1 ), and 2048 complex points with an acquisition time of 127 ms for 1 H (o 2 ). Repetition delays ranging from 1.2 (pk variants) to 1.7 s (S3L2 variants) were used between transients, with 128 scans per increment (total measuring times 36-49 h, respectively).