FIGURE 7.

Effect of Mg²⁺ and the 8U→C (U8C) mutation on reconstitution of the hmtRNA^Met from the D-half- and T-half-molecules. A, D-half-molecule (A₁-C₃₂, pink) and T-half-molecule (A₃₃-A₇₆, blue) were chemically synthesized to include Ψ₂₇ and Ψ₅₀. B, summary of the efficiency of reconstituting hmtRNA^Met from the normal (purple) and mutated (green) D-half- and T-half-molecules in the presence of Mg²⁺. C, titration of the U8 D-half-molecule (D) with increasing concentrations of the T-half-molecule (T) at 3 mm Mg²⁺ showing the formation of two complexes (C₁ and C₂). D, titration of the U8 D-half-molecule with increasing concentrations of the T-half-molecule in the absence of Mg²⁺ showing that no complex is formed. E, titration of the 8U→C D-half-molecule with increasing concentrations of the T-half-molecule at 3 mm Mg²⁺ showing that little complex is formed. F, titration of the 8U→C D-half-molecule with increasing concentrations of the T-half-molecule in the absence of Mg²⁺ showing that no complex is formed. The 1st lane labeled wt D/T is a positive control showing the formation of the complex between the normal U8 D-half-molecule (37.6 μm) and the normal T-half-molecule (37.1 μm). The lane labeled D_m indicates the migration of the 8U→C mutated D-half-molecule alone, whereas the T lane indicates the migration of the normal T-half-molecule.