JBC, Vol. 252, Issue 8, 2551-2559, Apr, 1977

Bisulfite-induced C changed to U transitions in yeast alanine tRNA

O. S. Bhanot and R. W. Chambers

The reaction of yeast tRNAAla1ab with NaHSO3 at 25 degrees and pH 5.8 has been studied. Five reactive residues have been located. Four of these (C-17 in Loop I, C-36 in the anticodon, C-74 and C-75 near the acceptor end) react to the same extent (42%) under the conditions of the experiment. The other (C-72 in the first base pair of the acceptor stem) reacts much more slowly (8%). No other changes were detected, but kinetic data suggest two or more additional residues may react very slowly. The C changed to U change in the anticodon (igc changed to igu) is a missense change (Ala changed to Thr). Both mechanistic considerations and experimental data from the literature show that HSO3--induced deamination of cytosine residues occurs only at unstacked residues. The quantitative changes for tRNAAla indicate that the stacking lifetimes of C-17, C-36, C-74, and C-75 are about equal. All other cytidine residues are much more tightly stacked. These results are consistent with the folded cloverleaf models that have been proposed from x-ray diffraction studies of yeast tRNAPhe. Residues 48 and 56, which are in single-stranded regions in the unfolded cloverleaf structure, do not react suggesting that they are tightly stacked in solution under the conditions of this experiment. The data also indicate that the anticodon loop is flexible in solution.
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