The Angle between the Anticodon and Aminoacyl Acceptor Stems of Yeast tRNAPhe Is Strongly Modulated by Magnesium Ions

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Abstract
Many tRNAs undergo tertiary folding transitions at temperatures well below the main thermally induced (hyperchromic) transition. Such transitions are essentially isochromic and isoenthalpic and display an absolute requirement for divalent cations; however, the nature of the structural transition is not known for any tRNA. Using a combination of transient electric birefringence (TEB) and gel electrophoretic measurements, we have characterized the influence of magnesium ions on the apparent angle between the anticodon and acceptor stems of a yeast tRNAPhe construct. TEB is a particularly sensitive method for quantifying the bends introduced in RNA by various nonhelix elements. In the current instance, the tRNA construct comprises an unmodified tRNAPhe molecule in which the anticodon and acceptor stems have been extended by ∼70 bp to more effectively “report” the interstem angles. Upon the addition of sub-millimolar concentrations of magnesium ions, the tRNA core undergoes a substantial rearrangement in tertiary structure, passing from an open form with an apparent interstem angle of ∼150° to a conformation with an interstem angle of ∼70° (200 μM Mg2+). Further addition of magnesium ions results in a minor adjustment of the apparent interstem angle to ∼80−90°, in line with earlier results. Finally, the magnesium-induced structural transition is essentially isochromic, in agreement with previous observations with native tRNAs. The current results suggest that changes in local divalent ion concentration in the ribosome could profoundly affect the global conformations of tRNAs during the translation cycle.