The methylthio group (ms2) of N6-(4-hydroxyisopentenyl)-2-methylthioadenosine (ms2io6A) present next to the anticodon contributes to the decoding efficiency of the tRNA

Abstract

A Salmonella typhimurium LT2 mutant which harbors a mutation (miaB2508::Tn10dCm) that results in a reduction in the activities of the amber suppressors supF30 (tRNA CUA Tyr ), supD10 (tRNA CUA Ser ), and supJ60 (tRNA CUA Leu ) was isolated. The mutant was deficient in the methylthio group (ms A37. The mutation miaB2508::Tn10dCm was 60% linked to the nag gene (min 15) and 40% linked to the fur gene and is located counterclockwise from both of these genes. The growth rates of the mutant in four growth media did not significantly deviate from those of a wild-type strain. The polypeptide chain elongation rate was also unaffected in the mutant. However, the miaB2508::Tn10dCm mutation rendered the cell more resistant or sensitive, compared with a wild-type cell, to several amino acid analogs, suggesting that this mutation influences the regulation of several amino acid biosynthetic operons. The efficiencies of the aforementioned amber suppressors were decreased to as low as 16%, depending on the suppressor and the codon context monitored, demonstrating that the ms 2 group of ms 2 io 6

Regulation of Substrate Recognition by the MiaA tRNA Prenyltransferase Modification Enzyme of Escherichia coliK-12

“…3, lane 4). The yield of His 6 -MiaA from 960 ml of bacterial culture was 6 mg with a specific activity of 700 nmol of i 6 A formed per min per mg of protein using synthetic tRNA Phe (wt) as a substrate (see below, and see "Experimental Procedures"). The His 6 -affinity tag could be completely cleaved from the fusion protein by thrombin protease (Fig.…”

“…In E. coli, the i 6 A37 tRNA modification is further methylthiolated by the action of the MiaB and MiaC enzyme activities to form ms 2 i 6 A37 ( Fig. 1), except for tRNA Sec (6). ms 2 i 6 A37-modified tRNA species read codons starting with U residues and include tRNA Phe , tRNA Trp , tRNA Tyr (I and II), tRNA Cys , tRNA Leu (IV and V), and tRNA Ser (II and III) but not tRNA Ser (I and V) (7,8).…”

MiaB Protein from Thermotoga maritima

“…Spectra were recorded digitally using photon-counting electronics, and improvements in signal-to-noise were achieved by signal averaging multiple scans. Band positions were calibrated using the excitation frequency and CCl 4 and are accurate to Ϯ1 cm Ϫ1 . Lines from a Coherent Sabre 10-W argon ion laser were used for excitation, and plasma lines were removed using a Pellin Broca prism premonochromator.…”

“…Changes in the degree of tRNA modification have been invoked as a possible global regulatory mechanism by which cells respond to certain environmental stresses (3). The importance of these modifications is well demonstrated by the fact that as much as 1% of the bacterial genome is devoted to tRNA modification (4). Modified bases are located throughout the tRNA molecule, but the greatest variety is found in the anticodon loop.…”

Disruption of MiaA provides insights into the regulation of phenazine biosynthesis under suboptimal growth conditions in Pseudomonas chlororaphis 30-84

“…In Escherichia coli, this tRNA prenyltransferase enzyme catalyses the addition of a D 2 -isopentenyl group from dimethylallyl diphosphate to the N 6 -nitrogen of adenosine in position 37 of specific tRNA species (i 6 A-37) [24,25]. After i 6 A-37 is formed by MiaA, the tRNA is further methylthiolated by the action of the MiaB (thiolation) and MiaC (methyl transfer) enzymes to form 2-methylthio-N 6 -(D 2 -isopentenyl) adenosine (ms 2 i 6 A-37) [26,27]. The tRNAs requiring MiaA-mediated modification recognize UXX codons, which includes all codons encoding Trp, Cys, Tyr and Phe, four of six tRNAs encoding Ser and two of six encoding Leu [25,[28][29][30].…”