ISOLATION OF THE GENE (MIAE) ENCODING THE HYDROXYLASE INVOLVED IN THESYNTHESIS OF 2-METHYLTHIO-CIS-RIBOZEATIN IN TRANSFER-RNA OF SALMONELLA-TYPHIMURIUM AND CHARACTERIZATION OF MUTANTS

The modified nucleoside 2-methylthio-N-6-isopentenyl adenosine (ms2i6A ) is present at position 37 (3' of the anticodon) of tRNAs that read c od ns beginning with U except tRNA(I,V)SER in Escherichia coli. Salmon ella typhimurium 2-methylthio-cis-ribozeatin (ms2io6A) is found in tRN A, probably in the corresponding species that have ms2i6A in E. coli. The gene (miaE) for the tRNA(ms2io6A)hydroxylase of S. typhimurium was isolated by complementation in E. coli. The miaE gene was localized c lose to the argI gene at min 99 of the S. typhimurium chromosomal map. Its DNA sequence and transcription pattern together with complementat ion studies revealed that the miaE gene is the second gene of a dicist ronic operon. Southern blot analysis showed that the miaE gene is abse nt in E. coli, a finding consistent with the absence of the hydroxylat ed derivative of ms2i6A in this species. Mutants of S. typhimurium whi ch have MudJ inserted in the miaE gene and which, consequently, are bl ocked in the ms2i6A hydroxylation reaction were isolated. Unexpectedly , such mutants cannot utilize the citric acid cycle intermediates mala te, fumarate, and succinate as carbon sources.