Structural elements that contribute to an unusual tertiary interaction in a transfer RNA.

Abstract

Transfer RNAs (tRNAs) contain a set of defined tertiary hydrogen-bonding interactions that are established between conserved and semiconserved nucleotides. Although the crystal structures of tRNAs describe each of the tertiary interactions in detailed molecular terms, little is known about the underlying structural parameters that stabilize the tertiary interactions. Escherichia coli (E. coli) tRNA(Cys) has an unusual tertiary interaction between G15 in the dihydrouridine (D) loop and G48 in the variable loop that is critical for cysteine aminoacylation. All other tRNAs have a purine 15 and a complementary pyrimidine 48 that establish a tertiary interaction known as the Levitt base pair [Levitt, M. (1969) Nature 224, 759-763; Klug et al. (1974) J. Mol. Biol. 89, 511-516]. In this study, the G15.G48 tertiary interaction in E. coli tRNA(Cys) was used to investigate the structural elements that contribute to its variation from the Levitt base pair. Analysis with chemical probes showed that substitution of U21 with A21 in the D loop and formation of a Watson-Crick base pair between nucleotides 13 and 22 in the D stem switch the hydrogen-pairing of G15.G48 to a Levitt-like G15.G48 base pair. This switch was accompanied by a decrease of the catalytic efficiency of aminoacylation by 2 orders of magnitude. In contrast, insertion of additional nucleotides in the D or variable loops had little effect.(ABSTRACT TRUNCATED AT 250 WORDS)