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Journal of Bacteriology, December 1999, p. 7256-7265, Vol. 181, No. 23
0021-9193/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.

Identification of the miaB Gene, Involved in Methylthiolation of Isopentenylated A37 Derivatives in the tRNA of Salmonella typhimurium and Escherichia coli

Birgitta Esberg,1 Hon-Chiu Eastwood Leung,2 Ho-Ching Tiffany Tsui,2,dagger Glenn R. Björk,1,* and Malcolm E. Winkler2,dagger

Department of Microbiology, Umeå University, S-901 87 Umeå, Sweden,1 and Department of Microbiology and Molecular Genetics, University of Texas, Houston, Medical School, Houston, Texas 77030-15012

Received 2 June 1999/Accepted 24 September 1999

The tRNA of the miaB2508::Tn10dCm mutant of Salmonella typhimurium is deficient in the methylthio group of the modified nucleoside N6-(4-hydroxyisopentenyl)-2-methylthioadenosine (ms2io6A37). By sequencing, we found that the Tn10dCm of this strain had been inserted into the f474 (yleA) open reading frame, which is located close to the nag locus in both S. typhimurium and Escherichia coli. By complementation of the miaB2508::Tn10dCm mutation with a minimal subcloned f474 fragment, we showed that f474 could be identified as the miaB gene, which is transcribed in the counterclockwise direction on the bacterial chromosome. Transcriptional studies revealed two promoters upstream of miaB in E. coli and S. typhimurium. A Rho-independent terminator was identified downstream of the miaB gene, at which the majority (96%) of the miaB transcripts terminate in E. coli, showing that the miaB gene is part of a monocistronic operon. A highly conserved motif with three cysteine residues was present in MiaB. This motif resembles iron-binding sites in other proteins. Only a weak similarity to an AdoMet-binding site was found, favoring the idea that the MiaB protein is involved in the thiolation step and not in the methylating reaction of ms2i(o)6A37 formation.

* Corresponding author. Mailing address: Department of Microbiology, Umeå University, S-901 87 Umeå, Sweden. Phone: 46-90-7856756. Fax: 46-90-772630. E-mail: glenn.bjork{at}micro.umu.se.

dagger Present address: Lilly Research Laboratories, Indianapolis, IN 46285.

Journal of Bacteriology, December 1999, p. 7256-7265, Vol. 181, No. 23
0021-9193/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.


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