Nucleotides of tRNA (Glu) involved in recognition by barley chloroplast glutamyl-tRNA synthetase and glutamyl-tRNA reductase

Robert D. Willows*, C. Gamini Kannangara, Bo Pontoppidan

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

16 Citations (Scopus)

Abstract

The biosynthesis of δ-aminolevulinate (ALA), via the C-5 pathway, requires tRNA(Glu) as a cofactor for the glutamyl tRNA(Glu) synthetase and the glutamyl tRNA(Glu) reductase which are the first two enzymes in this three step pathway. These two enzymes form a ternary complex with the tRNA(Glu) inChlamydomonas reinhardtii suggesting that the recognition elements on the tRNA cofactor are different for each enzyme. Chemical modification and comparative studies with tRNA(Glu)s from a number of species were used to determine the nucleotides involved in the recognition of the barley chloroplast tRNA(Glu) by the barley enzymes. The barley chloroplast tRNA(Glu) is chemically modified both before and after ligation to glutamate with monobromobimane or CNBr. The chemically modified tRNA(Glu) is a poor substrate for the glutamyl-tRNA synthetase and the chemically modified glutamyl-tRNA(Glu) is used as a substrate for glutamyl-tRNA(Glu) reductase. The tRNA(Glu) from the chloroplasts of barley,Chlamydomonas reinhardtii, tobacco, cucumber, wheat and spinach and tRNA(Glu) fromSynechocystis PCC6803, Escherichia coli, barley germ and bakers yeast and the barley chloroplast tRNA(Gln) are all effective substrates for the barley chloroplast glutamyl-tRNA synthetase. A comparison of the sequences of these tRNAs shows 19 conserved bases and five of these bases, G10, A26, U34, U35 and A37 are suggested as recognition elements of barley glutamyl tRNA(Glu) synthetase by assuming a similar binding orientation as in the crystal structure of theE. coli tRNA(Gln) GlnRS complex. The glutamyl-tRNA(Glu) fromE. coli, bakers yeast and bakers germ and the barley chloroplast glutamyl-tRNA(Gln) are not effective substrates for the barley chloroplast glutamyl-tRNA(Glu) reductase. A comparison of the sequences of these four tRNA species with the sequences of the tRNA(Glu) species that can be used as substrate by the glutamyl-tRNA(Glu) reductase yields seven common differences in the primary sequence. These 7 nucleotides, A7-U66, U29-A41, A53-U61, and U72 are expected to be required for recognition by the barley chloroplast glutamyl-tRNA(Glu) reductase.

Original languageEnglish
Pages (from-to)228-234
Number of pages7
JournalBBA - Gene Structure and Expression
Volume1263
Issue number3
DOIs
Publication statusPublished - 19 Sep 1995
Externally publishedYes

Keywords

  • (Barley)
  • Aminolevulinic acid
  • Chlorophyll biosynthesis
  • Glutamate
  • Glutamyl tRNA reductase
  • Glutamyl tRNA synthetase
  • tRNA identity

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