Assignment of imino proton signals of G-C base pairs and magnesium ion binding: an NMR study of bovine mitochondrial tRNA(SerGCU) lacking the entire D arm.

Abstract

The mammalian mitochondrial tRNA(SerGCU) (mt tRNA[SerGCU]) has a unique structure in that it lacks the whole D arm. To elucidate its higher-order structure, we synthesized unmodified bovine mt tRNA(SerGCU) using T7 RNA polymerase and measured its 1H-NMR spectrum in the imino proton region. Although the imino proton signals heavily overlapped, we succeeded in assigning all the seven imino proton signals of the G-C base pairs by a combination of base replacement and 15N-labeling of the G residues of a whole tRNA molecule or of the 3'-half fragment. The results indicate that the tRNA possesses the secondary structure that has been supposed on the basis of biochemical studies. Analysis of the effect of the magnesium concentration on the G-C pairs suggests that the acceptor and T stems do not form a co-axial helix, and that the core region of the tRNA does not interact with magnesium ions. These features are significantly different from those of canonical tRNAs. Despite this, it is very likely that the tRNA as a whole takes a nearly L-shape tertiary structure.