Functional Role of the Prokaryotic Proline-tRNA Synthetase Insertion Domain in Amino Acid Editing

Wong, Fai-Chu; Beuning, Penny J.; Nagan, Maria C.; Shiba, Kiyotaka; Musier‐Forsyth, Karin

doi:10.1021/bi012178j

Cited by 70 publications

(122 citation statements)

References 45 publications

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“…The highly conserved nature of the lysine residue that aligns with K279 of Ec ProRS is striking. This residue was previously shown to be critical for posttransfer editing of both Ec ProRS (16) and Hi YbaK (29), and we confirmed in this work that it also plays a critical role in editing by the EC/SC chimera. Two other residues are highly conserved among these domains: K284 and D350 (Ec ProRS numbering).…”

Section: Discussionsupporting

confidence: 88%

“…CP1 is present in all extant species and is believed to be of ancient origin (14). The class II synthetases, ProRS (15,16), alanyl-tRNA synthetase (AlaRS) (17,18), threonyl-tRNA synthetase (ThrRS) (19,20), and phenylalanyl-tRNA synthetase (21), contain a variety of editing domains that are unrelated to CP1. In the case of AlaRS, phylogenetic analysis suggested that, as for class I enzymes, the editing domain is present throughout evolution and co-evolved with the aminoacylation active site (18); it is thus believed to be of ancient origin.…”

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confidence: 99%

“…The unique ProRS editing domain is found in at least three different structural contexts: as an insertion (INS) between motifs 2 and 3 of the aminoacylation active site of bacterial enzymes (16,25,26), as an N-terminal extension in ProRSs of lower eukaryotes (27,28), and as a free-standing editing module (25,28). A highly conserved lysine (K279 in Ec ProRS) found in most INS domain homologs is critical for posttransfer editing function (16,29).…”

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confidence: 99%

“…A highly conserved lysine (K279 in Ec ProRS) found in most INS domain homologs is critical for posttransfer editing function (16,29). Archaeal and higher eukaryotic ProRSs lack an INS-like editing domain (Fig.…”

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confidence: 99%

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Restoring species-specific posttransfer editing activity to a synthetase with a defunct editing domain

SternJohn

Hati

Siliciano

et al. 2007

Proc. Natl. Acad. Sci. U.S.A.

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In contrast, the isolated INS domain displays only weak editing activity and lacks tRNA sequence specificity. These results emphasize the modular nature of synthetase editing active sites and demonstrate how in evolution, a weak editing activity can be converted to a more robust state through fusion to the body of a synthetase. In this manner, a single editing module can be distributed to different synthetases, and simultaneously acquire specificity and enhanced activity.Prolyl-tRNA synthetase ͉ Saccharomyces cerevisiae

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Section: Discussionsupporting

confidence: 88%

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confidence: 99%

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confidence: 99%

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confidence: 99%

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Restoring species-specific posttransfer editing activity to a synthetase with a defunct editing domain

SternJohn

Hati

Siliciano

et al. 2007

Proc. Natl. Acad. Sci. U.S.A.

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“…The second pathway is designated posttransfer editing. In this instance, the misactivated amino acid is transferred to the 3Ј end of tRNA to create a mischarged tRNA [such as Val-tRNA Ile (32), Ser-tRNA Thr (33), Ile-tRNA Phe (34), Ile-tRNA Leu (35), Ala-tRNA Pro (36), and Gly-tRNA Ala (3)]. The mischarged aminoacyl group is then cleared at the editing center.…”

mentioning

confidence: 99%

Interstice mutations that block site-to-site translocation of a misactivated amino acid bound to a class I tRNA synthetase

Bishop

Beebe

Schimmel

2003

Proc. Natl. Acad. Sci. U.S.A.

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Class I aminoacyl-tRNA synthetases catalyze editing reactions that prevent ambiguity from entering the genetic code. Misactivated amino acids are translocated in cis from the active site for aminoacylation to the center for editing, located Ϸ30 Å away. Mutational analysis has functionally separated the two sites by creating mutations that disrupt the catalytic center for editing but not for aminoacylation and vice versa. What is not known is whether translocation per se can be disrupted without an effect on either catalytic center. Here we describe mutations in a presumptive ''hinge region'' of isoleucyl-tRNA synthetase that is situated between the two sites. Interstice mutations had little or no effect on either catalytic center. In contrast, the same specific mutations disrupted translocation. Thus, with these mutations all three functions, translocation, catalysis of aminoacylation, and editing, have been mutationally separated. The results are consistent with translocation involving a hinge-region conformational shift that does not perturb the two catalytic centers.C onsiderable evidence supports the concept that the universal tree of life could not be generated without the editing activities of aminoacyl-tRNA synthetases (1-3). These ancient proteins catalyze aminoacylation reactions that are the basis of the genetic code, that is, the matching of amino acids with specific nucleotide triplets imbedded in transfer RNAs (4-8). Inherent limitations to the capacity of active sites to discriminate between closely similar amino acids (9) was compensated by the introduction of a second active center where misactivated amino acids are cleared (10-15). Without this active site for editing, ambiguity is introduced into the code as demonstrated by the extensive misincorporation of amino acids into cellular proteins when editing is disrupted (1).Class I aminoacyl-tRNA synthetases such as isoleucyl-, valyl-, and leucyl-tRNA synthetase are characterized by a catalytic center based on a Rossmann nucleotide-binding fold of alternating ␤-strands and ␣-helices (6, 16-21). The fold is split by an insertion known as connective polypeptide 1 (CP1) that joins one half of the catalytic domain to the other (12,(21)(22)(23). This insertion contains the active site for editing, located Ϸ30 Å from the catalytic center for aminoacylation (11-13, 15, 21, 24). The editing reactions require specific tRNAs as cofactors (25,26). The role of the tRNA is to trigger translocation of the misactivated residue from the catalytic center to the active site for editing (27)(28)(29).Amino acids are condensed with ATP to form aminoacyl adenylates bound to their cognate tRNA synthetases in the first step of protein synthesis. Occasional errors lead to activation of the wrong amino acid such as activation of valine by isoleucyltRNA synthetase (IleRS) or activation of threonine by valyltRNA synthetase (10, 30). Editing proceeds through two distinct pathways (Fig. 1). One is designated as the pretransfer step (10, 31). Here the misactivated amino acid (i...

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tRNA synthetase: tRNA aminoacylation and beyond

2014

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The aminoacyl-tRNA synthetases are prominently known for their classic function in the first step of protein synthesis, where they bear the responsibility of setting the genetic code. Each enzyme is exquisitely adapted to covalently link a single standard amino acid to its cognate set of tRNA isoacceptors. These ancient enzymes have evolved idiosyncratically to host alternate activities that go far beyond their aminoacylation role and impact a wide range of other metabolic pathways and cell signaling processes. The family of aminoacyl-tRNA synthetases have also been suggested as a remarkable scaffold to incorporate new domains that would drive evolution and the emergence of new organisms with more complex function. Because they are essential, the tRNA synthetases have served as pharmaceutical targets for drug and antibiotic development. The recent unfolding of novel important functions for this family of proteins offers new and promising pathways for therapeutic development to treat diverse human diseases.

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Functional Role of the Prokaryotic Proline-tRNA Synthetase Insertion Domain in Amino Acid Editing

Cited by 70 publications

References 45 publications

Restoring species-specific posttransfer editing activity to a synthetase with a defunct editing domain

Restoring species-specific posttransfer editing activity to a synthetase with a defunct editing domain

Interstice mutations that block site-to-site translocation of a misactivated amino acid bound to a class I tRNA synthetase

tRNA synthetase: tRNA aminoacylation and beyond

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