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. 2011 Jan 6;108(4):1308–1313. doi: 10.1073/pnas.1017085108

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Fig. 3. — The C terminus is required for efficient aminoacylation of a mutant . (A) lhp1Δ cells and trr4-1 lhp1Δ cells containing the indicated plasmids and pMETLHP1 were grown without methionine and switched to 2 mM methionine media at time 0. At intervals, RNA was extracted and subjected to Northern blotting to detect . As a loading control, the blot was reprobed to detect . (B) To separate aminoacylated and deacylated tRNAs, RNA was extracted under acidic conditions, fractionated in acidic polyacrylamide gels, and subjected to Northern blotting to detect and . Lane 1, deacylated tRNA from lhp1Δ cells. As described (11), both the charged and uncharged forms of trr4-1 migrate differently in these gels from the wild-type tRNA, consistent with adoption of an altered conformation by the mutant tRNA. (C) Quantification of the fraction of charged at intervals after methionine addition. Data from three independent experiments are graphed. Error bars are not visible for points where the S.E.M. was ≤ 1%.

Inline graphic — The C terminus is required for efficient aminoacylation of a mutant . (A) lhp1Δ cells and trr4-1 lhp1Δ cells containing the indicated plasmids and pMETLHP1 were grown without methionine and switched to 2 mM methionine media at time 0. At intervals, RNA was extracted and subjected to Northern blotting to detect . As a loading control, the blot was reprobed to detect . (B) To separate aminoacylated and deacylated tRNAs, RNA was extracted under acidic conditions, fractionated in acidic polyacrylamide gels, and subjected to Northern blotting to detect and . Lane 1, deacylated tRNA from lhp1Δ cells. As described (11), both the charged and uncharged forms of trr4-1 migrate differently in these gels from the wild-type tRNA, consistent with adoption of an altered conformation by the mutant tRNA. (C) Quantification of the fraction of charged at intervals after methionine addition. Data from three independent experiments are graphed. Error bars are not visible for points where the S.E.M. was ≤ 1%.