Microhelix aminoacylation by a class I tRNA synthetase. Non-conserved base pairs required for specificity.
Nucleotides in tRNAs that are conserved among isoacceptors are typically considered as candidates for tRNA synthetase recognition, with less importance attached to non-conserved nucleotides. Although the anticodon is an important contributor to the identity of methionine tRNAs, the class I methionine tRNA synthetase aminoacylates microhelices with high specificity. The microhelix substrates are comprised of as few as the 1st 4 base pairs of the acceptor stems of the elongator and initiator methionine tRNAs. For these two tRNAs, only the central 2:71 and 3:70 base pairs are common to the 1st 4 acceptor stem base pairs. We show here that, although the flanking 4:69 base pair is not conserved, a particular substitution at this position substantially reduces the gel electrophoresis-detected aminoacylation of an acceptor stem substrate that has the conserved 2:71 and 3:70 base pairs. Although the two methionine tRNAs have either U:A or G:C at position 4:69, substitution with C:G reduces charging of 9- or 4-base pair substrates that recreate part or all of the acceptor stem of a methionine tRNA. This effect is sufficient for methionine tRNA synthetase to discriminate between the closely related methionine and isoleucine tRNA acceptor stems. The ability to distinguish G:C and U:A from C:G is contrary to a simple scheme for recognition of atoms in the RNA minor groove.
Martinis
S A
SA
Department of Biology, Massachusetts Institute of Technology, Cambridge 02139.
Schimmel
P
P
D000215:Acylation; D001482:Base Composition; D001483:Base Sequence; D003196:Computer Graphics; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D008718:Methionine-tRNA Ligase / Q000378:metabolism*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D012329:RNA, Bacterial; D012358:RNA, Transfer, Met / Q000378:metabolism; D013379:Substrate Specificity
D012329:RNA, Bacterial; D012358:RNA, Transfer, Met; D008718:Methionine-tRNA Ligase