Nuclear magnetic resonance of the base-pairing structure of the native and denatured conformers of Escherichia coli transfer RNA Trp

Abstract

The native and denatured forms of transfer RNA Trp from Escherichia coli have been studied by high-resolution (300 MHz) proton nuclear magnetic resonance spectroscopy. The low-field nuclear magnetic resonance spectrum indicates that there are 22 ± 2 base-pairs present in the native form, and the observed resonance positions can be accounted for in terms of 19 base-pairs from the cloverleaf secondary structure, one protected U, and two tertiary structure base-pairs including A 14-s 4U 8. The denatured conformer contains one or two less base-pairs than the native conformer, and the low-field spectrum of the denatured conformer can be accounted for in terms of a model which retains the amino acid acceptor stem, T ΨC stem, and the anticodon stem of the cloverleaf model, but which pairs some bases of the dihydrouridine stem with bases from the minor loop. The proposed model is consistent with the ease of interconversion of the native and denatured conformers, the pattern of kethoxalation of the native and denatured conformera, and the retention of a tertiary structure A 14-s 4U 8 base-pair in the denatured conformer. Alternative models of the denatured form are considered, but they do not adequately explain the observed nuclear magnetic resonance spectrum. The binding of chloroquine to the native conformer has also been studied. A comparison of the binding of ethidium bromide and chloroquine to E. coli tRNA Glu indicates that the two drugs have very similar binding properties and this suggests a possible mechanism for the chloroquine-induced renaturation of tRNA Trp.