Structural and kinetic bases for the recognition of tRNAtyr by tyrosyl-tRNA synthetase

Labouze, Eric; Bedouelle, Hugues

doi:10.1016/0022-2836(89)90317-3

Cited by 44 publications

(35 citation statements)

References 17 publications

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“…Notably, nine of the fifteen amino acids involved in stabilizing the transition state for the first step of the reaction in the B. stearothermophilus enzyme (11,40,41) are conserved in the human, M. jannaschii, and S. cerevisiae enzymes. In contrast, none of the eleven amino acids known to be involved in tRNA Tyr recognition (42)(43)(44)(45)(46)(47) are conserved between the human and B. stearothermophilus tyrosyl-tRNA synthetases, suggesting that in contrast to the mechanism for formation of the E⅐Tyr-AMP intermediate, tRNA Tyr recognition differs between eukaryotes, archaea, and bacteria. This is most apparent in the M. jannaschii amino acid sequence, which surprisingly is missing a substantial portion of the tRNA Tyr anticodon recognition domain (amino acids 330 -419 in the B. stearothermophilus enzyme).…”

Section: Resultsmentioning

confidence: 99%

Human Tyrosyl-tRNA Synthetase Shares Amino Acid Sequence Homology with a Putative Cytokine

Kleeman¹,

Wei²,

Simpson

et al. 1997

Journal of Biological Chemistry

116

102

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To test the hypothesis that tRNA Tyr recognition differs between bacterial and human tyrosyl-tRNA synthetases, we sequenced several clones identified as human tyrosyl-tRNA synthetase cDNAs by the Human Genome Project. We found that human tyrosyl-tRNA synthetase is composed of three domains: 1) an aminoterminal Rossmann fold domain that is responsible for formation of the activated E⅐Tyr-AMP intermediate and is conserved among bacteria, archeae, and eukaryotes; 2) a tRNA anticodon recognition domain that has not been conserved between bacteria and eukaryotes; and 3) a carboxyl-terminal domain that is unique to the human tyrosyl-tRNA synthetase and whose primary structure is 49% identical to the putative human cytokine endothelial monocyte-activating protein II, 50% identical to the carboxyl-terminal domain of methionyl-tRNA synthetase from Caenorhabditis elegans, and 43% identical to the carboxyl-terminal domain of Arc1p from Saccharomyces cerevisiae. The first two domains of the human tyrosyl-tRNA synthetase are 52, 36, and 16% identical to tyrosyl-tRNA synthetases from S. cerevisiae, Methanococcus jannaschii, and Bacillus stearothermophilus, respectively. Nine of fifteen amino acids known to be involved in the formation of the tyrosyl-adenylate complex in B. stearothermophilus are conserved across all of the organisms, whereas amino acids involved in the recognition of tRNA Tyr are not conserved. Kinetic analyses of recombinant human and B. stearothermophilus tyrosyl-tRNA synthetases expressed in Escherichia coli indicate that human tyrosyl-tRNA synthetase aminoacylates human but not B. stearothermophilus tRNA Tyr , and vice versa, supporting the original hypothesis. It is proposed that like endothelial monocyte-activating protein II and the carboxyl-terminal domain of Arc1p, the carboxyl-terminal domain of human tyrosyltRNA synthetase evolved from gene duplication of the carboxyl-terminal domain of methionyl-tRNA synthetase and may direct tRNA to the active site of the enzyme.Aminoacyl-tRNA synthetases catalyze the aminoacylation of tRNA by their cognate amino acid. For most aminoacyl-tRNA synthetases (E), tRNA aminoacylation can be separated into two steps: formation of a stable enzyme-bound aminoacyladenylate intermediate (E⅐AA-AMP, Equation 1), followed by transfer of the amino acid (AA) from the aminoacyl-adenylate intermediate to the 3Ј end of the tRNA substrate (Equation 2).The 20 different aminoacyl-tRNA synthetases fall into two distinct structural classes (1, 2). Class I aminoacyl-tRNA synthetases (of which tyrosyl-tRNA synthetase is a member) are characterized by a structurally well conserved amino-terminal Rossmann fold domain which contains the signature sequences "HIGH" and "KMSKS" (3, 4). In contrast, the carboxyl-terminal domains of class I aminoacyl-tRNA synthetases are structurally diverse suggesting that the primordial aminoacyl-tRNA synthetase consisted solely of the amino-terminal Rossmann fold (5-8). Both x-ray crystallography and site-directed mutagenesis of class I aminoacyl-tRNA synthet...

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Section: Resultsmentioning

confidence: 99%

Human Tyrosyl-tRNA Synthetase Shares Amino Acid Sequence Homology with a Putative Cytokine

Kleeman¹,

Wei²,

Simpson

et al. 1997

Journal of Biological Chemistry

116

102

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“…3). This position of tRNA, base 73, is designated the discriminator base (3) and is an important identity determinant for a number of tRNAs, including tRNATYr (27,37,38).…”

Section: Resultsmentioning

confidence: 99%

Interaction between the acceptor end of tRNA and the T box stimulates antitermination in the Bacillus subtilis tyrS gene: a new role for the discriminator base

1994

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The Bacillus subtilis tyrS gene is a member of a group of gram-positive aminoacyl-tRNA synthetase and amino acid biosynthesis genes which are regulated by transcription antitermination. Each gene in the group is specifically induced by limitation for the appropriate amino acid. This response is mediated by interaction of the cognate tRNA with the mRNA leader region to promote formation of an antiterminator structure. The tRNA interacts with the leader by codon-anticodon pairing at a position designated the specifier sequence which is upstream of the antiterminator. In this study, an additional site of possible contact between the tRNA and the leader was identified through covariation of leader mRNA and tRNA sequences. Mutations in the acceptor end of tRNAl',r could suppress mutations in the side bulge of the antiterminator, in a pattern consistent with base pairing. This base pairing may thereby directly affect the formation and/or function of the antiterminator. The discriminator position of the tRNA, an important identity determinant for a number of tRNAs, including tRNA"tF, was shown to act as a second specificity determinant for assuring response to the appropriate tRNA. Furthermore, overproduction of an unchargeable variant of tRNATYr resulted in antitermination in the absence of limitation for tyrosine, supporting the proposal that uncharged tRNA is the effector in this system.Most of the aminoacyl-tRNA synthetase genes identified to date in Bacillus subtilis are regulated by transcription antitermination (17,22,24,35). All of these genes exhibit very strong conservation of a set of primary-sequence and secondarystructure elements in their mRNA leader regions, upstream of the start of the coding region (17, 18). These elements, which include three stem-loop structures, a highly conserved 14-bp sequence designated the T box, and a factor-independent transcriptional terminator ( Fig. 1), are also found in certain amino acid biosynthesis genes and tRNA synthetase genes in Bacillus and other gram-positive species, including B. subtilis ilv-leu and cysE-cysS, Lactobacillus casei valS and trp, Lactococcus lactis trp and his, and Brevibacterium lactofermentum (and Corynebacterium glutamicum) argS-lysA, suggesting that this mechanism is widespread in gram-positive bacteria (18,22). For each gene that has been examined, readthrough of the leader region terminator is induced by limitation for the appropriate amino acid and not by general amino acid starvation (5,12,13,24,35). Analysis of the mechanism for the specific response of each gene to the cognate amino acid has been a major focus of attention in our laboratory.We noted that each leader region structure contains a triplet sequence corresponding to a codon specifying the appropriate amino acid, displayed at a specific position within a side bulge of the first stem-loop structure (17). Alteration of this triplet in the tyrS leader from a UAC tyrosine codon to a UUC phenylalanine codon was sufficient to switch the specificity of the response to amino acid limita...

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“…These variable features, including the size of the variable arm, nucleotide identity at position 73 (which is designated the discriminator base), and the structure of the acceptor stem, often serve in conjunction with the anticodon as identity elements for correct aminoacylation (Labouze and Bedouelle 1989;Sherman et al 1992a,b;Giege et al 1993;McClain et al 1999). We previously showed that substitutions in the variable arm, anticodon and position 73 of the tRNA, did not block antitermination activity, as long as base-pairing at the specifier sequence and antiterminator variable position (position 158 in glyQS) was maintained (Grundy et al 2000(Grundy et al , 2002.…”

Section: Discussionmentioning

confidence: 99%

tRNA requirements for glyQS antitermination: A new twist on tRNA

Yousef

Grundy²,

Henkin³

2003

RNA

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Transcription antitermination of the Bacillus subtilis glyQS gene, a member of the T box gene regulation family, can be induced during in vitro transcription in a minimal system using purified B. subtilis RNA polymerase by the addition of unmodified T7 RNA polymerase-transcribed tRNA Gly. Antitermination was previously shown to depend on base-pairing between the glyQS leader and the tRNA at the anticodon and acceptor ends. In this study, variants of tRNA Gly were generated to identify additional tRNA elements required for antitermination activity, and to determine the effect of structural changes in the tRNA. We find that additions to the 3 end of the tRNA blocked antitermination, in agreement with the prediction that uncharged tRNA is the effector in vivo, whereas insertion of 1 nucleotide between the acceptor stem and the 3 UCCA residues had no effect. Disruption of the D-loop/T-loop tertiary interaction inhibited antitermination function, as was previously demonstrated for tRNATyr -directed antitermination of the B. subtilis tyrS gene in vivo. Insertion of a single base pair in the anticodon stem was tolerated, whereas further insertions abolished antitermination. However, we find that major alterations in the length of the acceptor stem are tolerated, and the insertions exhibited a pattern of periodicity suggesting that there is face-of-the-helix dependence in the positioning of the unpaired UCCA residues at the 3 end of the tRNA for interaction with the antiterminator bulge and antitermination.

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Structural and kinetic bases for the recognition of tRNAtyr by tyrosyl-tRNA synthetase

Cited by 44 publications

References 17 publications

Human Tyrosyl-tRNA Synthetase Shares Amino Acid Sequence Homology with a Putative Cytokine

Human Tyrosyl-tRNA Synthetase Shares Amino Acid Sequence Homology with a Putative Cytokine

Interaction between the acceptor end of tRNA and the T box stimulates antitermination in the Bacillus subtilis tyrS gene: a new role for the discriminator base

tRNA requirements for glyQS antitermination: A new twist on tRNA

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