Transfer RNA recognition by the Escherichia coli delta2-isopentenyl-pyrophosphate:tRNA delta2-isopentenyl transferase: dependence on the anticodon arm structure

RNA. 1997 Jul;3(7):721-33.

Abstract

To elucidate the sequence elements required in the anticodon stem for the recognition of Escherichia coli tRNA(Ser) (GGA) by the E. coli isopentenyl-tRNA:A37 transferase (IPTT), which result in the conversion of A37 into isopentenylated i6A37, we have tested and characterized in vitro T7-runoff transcripts of 17 variants of E. coli tRNA(Ser)(GGA) and 7 other tRNAs from E. coli and yeast. Our results indicate that, instead of a stringent specific anticodon stem and loop sequence, the key feature required for the recognition of E. coli tRNAs by IPTT is the A36A37A38 sequence occurring within the seven-membered anticodon loop, and the retention of the standard helical structure and flexibility, especially in the proximal anticodon stem. The G30*U40 mismatch base pair close to the anticodon loop is strictly avoided. The frequent occurrence of a C-G base pair in the three stem locations closest to the loop (positions 29-41, 30-40 and 31-39) or the occurrence of even one such C-G base pair along with some other similarly less suited, but individually tolerated deviations can also totally abolish the A37 isopentenylation of tRNA. For the position 30-40, the G-C base pair is shown uniquely suited, whereas for the adjoining 29-41 stem location, a purine-pyrimidine base pair with pyrimidine on the 3'-side is strongly preferred. Retention of the overall 3D tRNA structure is favorable for isopentenylation and allows some tolerance of proximal stem sequence deviations. Our data suggest a recognition mode that implies the interaction of IPTT with the strictly conserved A36A37A38 sequence and the other functional groups located in the minor groove of the anticodon stem.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Alkyl and Aryl Transferases*
  • Anticodon / chemistry*
  • Anticodon / metabolism
  • Base Composition
  • Base Sequence
  • Conserved Sequence
  • Databases, Factual
  • Escherichia coli / enzymology*
  • Escherichia coli / genetics
  • Isopentenyladenosine / metabolism
  • Molecular Sequence Data
  • Mutation
  • Nucleic Acid Conformation
  • RNA, Transfer / chemistry
  • RNA, Transfer / genetics
  • RNA, Transfer / metabolism*
  • RNA, Transfer, Ser / chemistry
  • RNA, Transfer, Ser / genetics
  • RNA, Transfer, Ser / metabolism
  • Structure-Activity Relationship
  • Substrate Specificity
  • Transcription, Genetic
  • Transferases / metabolism*

Substances

  • Anticodon
  • RNA, Transfer, Ser
  • Isopentenyladenosine
  • RNA, Transfer
  • Transferases
  • Alkyl and Aryl Transferases
  • adenylate isopentenyltransferase