Effect of modifications on native state stability and
dynamics
of bmt tRNALeu. (A) Leucylation reactions of the bmt tRNALeuUAA transcript performed at different concentrations
of Mg2+. The tRNA was preincubated in 2.5 mM ATP, 10 mM
Leu, and the indicated concentrations of Mg2+ for 30 min
before initiating the reaction with the addition of 5 μM LARS2.
(B) Leucylation reaction of bmt tRNALeuUAA transcript
preincubated either in the absence or presence of 20 mM Mg2+, before simultaneously initiating the reaction with 5 μM LARS2
and readjusting the Mg2+ concentration to 20 mM Mg2+. Rate constants for the slow phases were 1.27 min–1 (Δ) and 1.42 min–1 (∇), and the Y-intercepts
were 0.53 and 0.86, respectively. The rate constants for the fast
phases were too fast to be measured by manual pipetting. (C) Leucylation
reactions of modified bmt tRNALeu isolated from cells,
performed as described for panel A. (D) Plot of the normalized fraction
of the native state as a function of Mg2+ concentration.
The half-maximum Mg2+ concentrations for folding to the
native state (Mg1/2) are 0.59 mM for modified tRNA and
2.23 mM for the unmodified transcript tRNA.