Alanine Transfer RNA Synthetase: Structure–Function Relationships and Molecular Recognition of Transfer RNA

Alanyl-tRNA synthetases (AlaRSs) from three domains of life predominantly rely on a single wobble base pair, G3-U70, of tRNA Ala as a major determinant. However, this base pair is divergent in human mitochondrial tRNA Ala , but instead with a translocated G5-U68. How human mitochondrial AlaRS (hmtAlaRS) recognizes tRNA Ala , in particular, in the acceptor stem region, remains unknown. In the present study, we found that hmtAlaRS is a monomer and recognizes mitochondrial tRNA Ala in a G3-U70-independent manner, requiring several elements in the acceptor stem. In addition, we found that hmtAlaRS misactivates noncognate Gly and catalyzes strong transfer RNA (tRNA)-independent pre-transfer editing for Gly. A completely conserved residue outside of the editing active site, Arg 663 , likely functions as a tRNA translocation determinant to facilitate tRNA entry into the editing domain during editing. Finally, we investigated the effects of the severe infantile-onset cardiomyopathy-associated R592W mutation of hmtAlaRS on the canonical enzymatic activities of hmtAlaRS. Overall, our results provide fundamental information about tRNA recognition and deepen our understanding of translational quality control mechanisms by hmtAlaRS.

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“…AlaRS is responsible for producing Ala-tRNA Ala (36). However, bacterial AlaRSs have been shown to misactivate noncognate Ser and Gly (37).…”

Section: Introductionmentioning

confidence: 99%

The G3-U70-independent tRNA recognition by human mitochondrial alanyl-tRNA synthetase

Zeng

Peng

et al. 2019

Nucleic Acids Research

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“…Aminoacyl-tRNA synthetases attach amino acids to their cognate tRNAs in protein biosynthesis − through an aminoacyl-adenylate intermediate. Analysis of the thermodynamics, dynamics, and kinetics of coupled ligand interaction with these enzymes is essential to understanding their catalytic mechanisms.…”

mentioning

confidence: 99%

Allosteric Interaction of Nucleotides and tRNA^ala with E. coli Alanyl-tRNA Synthetase

et al. 2011

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Alanyl-tRNA synthetase, a dimeric class 2 aminoacyl-tRNA synthetase, activates glycine and serine at significant rates. An editing activity hydrolyzes Gly-tRNA(ala) and Ser-tRNA(ala) to ensure fidelity of aminoacylation. Analytical ultracentrifugation demonstrates that the enzyme is predominately a dimer in solution. ATP binding to full length enzyme (ARS875) and to an N-terminal construct (ARS461) is endothermic (ΔH = 3-4 kcal mol(-1)) with stoichiometries of 1:1 for ARS461 and 2:1 for full-length dimer. Binding of aminoacyl-adenylate analogues, 5'-O-[N-(L-alanyl)sulfamoyl]adenosine (ASAd) and 5'-O-[N-(L-glycinyl)sulfamoyl]adenosine (GSAd), are exothermic; ASAd exhibits a large negative heat capacity change (ΔC(p) = 0.48 kcal mol(-1) K(-1)). Modification of alanyl-tRNA synthetase with periodate-oxidized tRNA(ala) (otRNA(ala)) generates multiple, covalent, enzyme-tRNA(ala) products. The distribution of these products is altered by ATP, ATP and alanine, and aminoacyl-adenylate analogues (ASAd and GSAd). Alanyl-tRNA synthetase was modified with otRNA(ala), and tRNA-peptides from tryptic digests were purified by ion exchange chromatography. Six peptides linked through a cyclic dehydromoropholino structure at the 3'-end of tRNA(ala) were sequenced by mass spectrometry. One site lies in the N-terminal adenylate synthesis domain (residue 74), two lie in the opening to the editing site (residues 526 and 585), and three (residues 637, 639, and 648) lie on the back side of the editing domain. At least one additional modification site was inferred from analysis of modification of ARS461. The location of the sites modified by otRNA(ala) suggests that there are multiple modes of interaction of tRNA(ala) with the enzyme, whose distribution is influenced by occupation of the ATP binding site.

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Comparison of dissimilarity patterns of E coli, yeast and mammalian tRNAs

Steinberg

Kisselev

1992

Biochimie

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Alanine Transfer RNA Synthetase: Structure–Function Relationships and Molecular Recognition of Transfer RNA

Cited by 7 publications

References 94 publications

The G3-U70-independent tRNA recognition by human mitochondrial alanyl-tRNA synthetase

The G3-U70-independent tRNA recognition by human mitochondrial alanyl-tRNA synthetase

Allosteric Interaction of Nucleotides and tRNA^ala with E. coli Alanyl-tRNA Synthetase

Comparison of dissimilarity patterns of E coli, yeast and mammalian tRNAs

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Alanine Transfer RNA Synthetase: Structure–Function Relationships and Molecular Recognition of Transfer RNA

Cited by 7 publications

References 94 publications

The G3-U70-independent tRNA recognition by human mitochondrial alanyl-tRNA synthetase

The G3-U70-independent tRNA recognition by human mitochondrial alanyl-tRNA synthetase

Allosteric Interaction of Nucleotides and tRNAala with E. coli Alanyl-tRNA Synthetase

Comparison of dissimilarity patterns of E coli, yeast and mammalian tRNAs

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Allosteric Interaction of Nucleotides and tRNA^ala with E. coli Alanyl-tRNA Synthetase