Fig. 6.
Pus7 is more active at elevated temperatures on substrates with UGUAR sequences predicted to be in secondary structures. (A) Time courses of Ψ incorporation into ST1 by wild-type Pus7 at varying temperatures (●: 30 °C; □: 37 °C; ▲: 42 °C; and ○: 50 °C). The single-turnover kobs values for these reactions are reported in SI Appendix, Fig. S19. (B) Differences in the stoichiometry of Ψ incorporation at 30 °C and 37 °C in full-length RNA substrates measured by CLAP. The level of Ψ addition is generally enhanced at sites that were only detectable under heat shock in Schwartz et al. (24). (C) Representative CLAP gel of BET2 pseudouridylated target site from total RNA purified from BY4741 yeast pus7Δ::kanMX. Black arrow denotes the truncated, pseudouridylated product. The upper band is the unmodified, full-length product.