Leucyl-tRNA synthetase from the ancestral bacterium Aquifex aeolicus contains relics of synthetase evolution

Zhao, Mengchen; Zhu, Bin; Hao, Rui; Xu, Mingang; Eriani, Gilbert; Wang, En‐Duo

doi:10.1038/sj.emboj.7600618

Cited by 40 publications

(83 citation statements)

References 65 publications

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“…The first coarse sieve, an aminoacylation active site that is located in an ancient canonical core of the aaRS, is responsible for activating amino acids. A second fine sieve, a hydrolytic active site, has been clearly demonstrated to correct the enzyme's mistakes in a number of synthetase systems (27)(28)(29)(30)(31)(32)(33)(34)(35).…”

Section: Introduction: Aminoacyl-trna Synthetase Fidelitymentioning

confidence: 99%

In vitro assays for the determination of aminoacyl-tRNA synthetase editing activity

Splan

Musier‐Forsyth

Boniecki

et al. 2008

Methods

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Aminoacyl-tRNA synthetases are essential enzymes that help to ensure the fidelity of protein translation by accurately aminoacylating (or "charging") specific tRNA substrates with cognate amino acids. Many synthetases have an additional catalytic activity to confer amino acid editing or proofreading. This activity relieves ambiguities during translation of the genetic code that result from one synthetase activating multiple amino acid substrates. In this review, we describe methods that have been developed for assaying both pre-and post-transfer editing activities. Pre-transfer editing is defined as hydrolysis of a misactivated aminoacyl-adenylate prior to transfer to the tRNA. This reaction has been reported to occur either in the aminoacylation active site or in a separate editing domain. Post-transfer editing refers to the hydrolysis reaction that cleaves the aminoacyl-ester linkage formed between the carbonyl carbon of the amino acid and the 2′ or 3′ hydroxyl group of the ribose on the terminal adenosine. Post-transfer editing takes place in a hydrolytic active site that is distinct from the site of amino acid activation. Here, we focus on methods for determination of steady-state reaction rates using editing assays developed for both classes of synthetases.

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Section: Introduction: Aminoacyl-trna Synthetase Fidelitymentioning

confidence: 99%

In vitro assays for the determination of aminoacyl-tRNA synthetase editing activity

Splan

Musier‐Forsyth

Boniecki

et al. 2008

Methods

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“…Examination of the IleRS, ValRS, and LeuRS CP1 domains as free-standing fragments showed that they possess hydrolytic activity toward misacylated tRNA (11)(12)(13). Analogs of misacylated tRNA were shown to bind to a threonine-rich cleft in the CP1 domains of IleRS and LeuRS, uncovering a proofreading site located 30 Å away from the synthetic site (14,15) and pinpointing specific amino acid residues that may participate in post-transfer editing.…”

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confidence: 99%

Partitioning of tRNA-dependent Editing between Pre- and Post-transfer Pathways in Class I Aminoacyl-tRNA Synthetases

Dulić

Cvetešić

Perona

et al. 2010

Journal of Biological Chemistry

175

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Hydrolytic editing activities are present in aminoacyl-tRNA synthetases possessing reduced amino acid discrimination in the synthetic reactions. Post-transfer hydrolysis of misacylated tRNA in class I editing enzymes occurs in a spatially separate domain inserted into the catalytic Rossmann fold, but the location and mechanisms of pre-transfer hydrolysis of misactivated amino acids have been uncertain. Here, we use novel kinetic approaches to distinguish among three models for pre-transfer editing by Escherichia coli isoleucyl-tRNA synthetase (IleRS). We demonstrate that tRNA-dependent hydrolysis of noncognate valyl-adenylate by IleRS is largely insensitive to mutations in the editing domain of the enzyme and that noncatalytic hydrolysis after release is too slow to account for the observed rate of clearing. Measurements of the microscopic rate constants for amino acid transfer to tRNA in IleRS and the related valyl-tRNA synthetase (ValRS) further suggest that pre-transfer editing in IleRS is an enzyme-catalyzed activity residing in the synthetic active site. In this model, the balance between pretransfer and post-transfer editing pathways is controlled by kinetic partitioning of the noncognate aminoacyl-adenylate. Rate constants for hydrolysis and transfer of a noncognate intermediate are roughly equal in IleRS, whereas in ValRS transfer to tRNA is 200-fold faster than hydrolysis. In consequence, editing by ValRS occurs nearly exclusively by post-transfer hydrolysis in the editing domain, whereas in IleRS both pre-and post-transfer editing are important. In both enzymes, the rates of amino acid transfer to tRNA are similar for cognate and noncognate aminoacyl-adenylates, providing a significant contrast with editing DNA polymerases.

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“…91 Proofreading to remove amino acids from noncognate tRNAs was likely to have been as necessary for A73 tRNAs as it is for A76 tRNAs. Given the abilities of separately expressed editing domains to deacylate A76 tRNAs, 152,153 and the existence of separate proteins for trans-editing in some cases, 92,93,137 similar editing proteins may have served the same purpose for A73 tRNAs. Hence, editing domains in aaRSs may have been carried over from the A73 tRNA period.…”

Section: Evolution Of Trnasmentioning

confidence: 99%

An alternative to the RNA World Hypothesis

Francis

2011

Trends Evol Biol

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The RNA world hypothesis for the origin of life is widely accepted in spite of the complexity of RNA synthesis. An alternative hypothesis is advanced for the origin and evolution of protein and nucleic acid synthesis. From an early stage synthesis of the evolutionary predecessors of nucleic acids and polypeptides was coupled. This evolved into an RNA replication process that used 3'-aminoacyl-NMP monomers (N= pyrimidine or purine) with a singlet coding system in which the four bases coded for glycine, alanine, valine and aspartic acid. Ribosomal protein synthesis (RPS) evolved from coupled replication by using tRNAs and separating fidelity checking and peptide bond formation functions into small and large ribosomal subunits. Continuity was maintained during the transition to the triplet process by using the same catalysts that aminoacylated NMPs to aminoacylate four different bases at the 3' ends of the original tRNAs. Four GNC codons used the central base to designate the same four amino acids that were coded in singlet replication. Triplet-coded protein synthesis had the advantage of producing multiple copies of protein from a single copy of RNA, and eventually replaced protein synthesis via singlet-coded replication. Evolution of the simple triplet-coded process into RPS is described.

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Leucyl-tRNA synthetase from the ancestral bacterium Aquifex aeolicus contains relics of synthetase evolution

Cited by 40 publications

References 65 publications

In vitro assays for the determination of aminoacyl-tRNA synthetase editing activity

In vitro assays for the determination of aminoacyl-tRNA synthetase editing activity

Partitioning of tRNA-dependent Editing between Pre- and Post-transfer Pathways in Class I Aminoacyl-tRNA Synthetases

An alternative to the RNA World Hypothesis

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