Crystal Structure of Tryptophanyl-tRNA Synthetase Complexed with Adenosine-5′ Tetraphosphate: Evidence for Distributed Use of Catalytic Binding Energy in Amino Acid Activation by Class I Aminoacyl-tRNA Synthetases

Aminoacyl-tRNA synthetases produce aminoacyl-tRNAs from the substrate tRNA and its cognate amino acid with the aid of ATP. Two types of glutamyl-tRNA synthetase (GluRS) have been discovered: discriminating GluRS (D-GluRS) and nondiscriminating GluRS (ND-GluRS). D-GluRS glutamylates tRNAGluonly, while ND-GluRS glutamylates both tRNAGluand tRNAGln. ND-GluRS produces the intermediate Glu-tRNAGln, which is converted to Gln-tRNAGlnby Glu-tRNAGlnamidotransferase. Two GluRS homologues fromThermotoga maritima, TM1875 and TM1351, have been biochemically characterized and it has been clarified that only TM1875 functions as an ND-GluRS. Furthermore, the crystal structure of theT. maritimaND-GluRS, TM1875, was determined in complex with a Glu-AMP analogue at 2.0 Å resolution. TheT. maritimaND-GluRS contains a characteristic structure in the connective-peptide domain, which is inserted into the catalytic Rossmann-fold domain. The glutamylation ability of tRNAGlnby ND-GluRS was measured in the presence of the bacterial Glu-tRNAGlnamidotransferase GatCAB. Interestingly, the glutamylation efficiency was not affected even in the presence of excess GatCAB. Therefore, GluRS avoids competition with GatCAB and glutamylates tRNAGln.

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Amino acid activation analysis of primitive aminoacyl-tRNA synthetases encoded by both strands of a single gene using the malachite green assay

Biosystems ◽

10.1016/j.biosystems.2021.104481 ◽

2021 ◽

pp. 104481

Author(s):

Kazaha Onodera ◽

Nana Suganuma ◽

Haruka Takano ◽

Yu Sugita ◽

Tomoko Shoji ◽

...

Keyword(s):

Amino Acid ◽

Activation Analysis ◽

Malachite Green ◽

Single Gene ◽

Acid Activation ◽

Trna Synthetases ◽

Amino Acid Activation ◽

Aminoacyl Trna Synthetases ◽

Malachite Green Assay

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Inhibitory mechanism of reveromycin A at the tRNA binding site of a class I synthetase

Nature Communications ◽

10.1038/s41467-021-21902-0 ◽

2021 ◽

Vol 12 (1) ◽

Author(s):

Bingyi Chen ◽

Siting Luo ◽

Songxuan Zhang ◽

Yingchen Ju ◽

Qiong Gu ◽

...

Keyword(s):

Binding Site ◽

Catalytic Domain ◽

Trna Synthetase ◽

Rational Drug Design ◽

Class I ◽

Binding Assays ◽

Trna Synthetases ◽

Aminoacyl Trna Synthetases ◽

Rational Drug ◽

Trna Binding

AbstractThe polyketide natural product reveromycin A (RM-A) exhibits antifungal, anticancer, anti-bone metastasis, anti-periodontitis and anti-osteoporosis activities by selectively inhibiting eukaryotic cytoplasmic isoleucyl-tRNA synthetase (IleRS). Herein, a co-crystal structure suggests that the RM-A molecule occupies the substrate tRNAIle binding site of Saccharomyces cerevisiae IleRS (ScIleRS), by partially mimicking the binding of tRNAIle. RM-A binding is facilitated by the copurified intermediate product isoleucyl-adenylate (Ile-AMP). The binding assays confirm that RM-A competes with tRNAIle while binding synergistically with l-isoleucine or intermediate analogue Ile-AMS to the aminoacylation pocket of ScIleRS. This study highlights that the vast tRNA binding site of the Rossmann-fold catalytic domain of class I aminoacyl-tRNA synthetases could be targeted by a small molecule. This finding will inform future rational drug design.

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Six Rossmannoid folds, including the Class I aminoacyl-tRNA synthetases, share a partial core with the anti-codon-binding domain of a Class II aminoacyl-tRNA synthetase

Bioinformatics ◽

10.1093/bioinformatics/btq039 ◽

2010 ◽

Vol 26 (6) ◽

pp. 709-714 ◽

Cited By ~ 28

Author(s):

S. Cammer ◽

C. W. Carter

Keyword(s):

Class Ii ◽

Trna Synthetase ◽

Binding Domain ◽

Class I ◽

Aminoacyl Trna Synthetase ◽

Trna Synthetases ◽

Aminoacyl Trna Synthetases

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Functional characterization of leucine-specific domain 1 from eukaryal and archaeal leucyl-tRNA synthetases

Biochemical Journal ◽

10.1042/bj20100235 ◽

2010 ◽

Vol 429 (3) ◽

pp. 505-513 ◽

Cited By ~ 5

Author(s):

Xiao-Long Zhou ◽

Meng Wang ◽

Min Tan ◽

Qian Huang ◽

Gilbert Eriani ◽

...

Keyword(s):

Functional Characterization ◽

Trna Synthetase ◽

Acid Activation ◽

Reaction Step ◽

Specific Domain ◽

Binding Strength ◽

Trna Synthetases ◽

Amino Acid Activation ◽

Common Domain

LeuRS (leucyl-tRNA synthetase) catalyses the esterification of tRNAsLeu with leucine. This family of enzymes is divided into prokaryotic and eukaryal/archaeal groups according to the presence and position of specific insertions and extensions. In the present study, we investigated the function of LSD1 (leucine-specific domain 1), which is naturally present in eukaryal/archaeal LeuRSs, but absent from prokaryotic LeuRSs. When mutated in their common domain, the eukaryal and archaeal LeuRSs exhibited defects in the first reaction step of amino acid activation with variations of leucine or ATP-binding strength, whereas the tRNA aminoacylation was moderately affected. When the eukaryal extension was mutated, severe tRNA charging defects were observed, suggesting that eukaryotes evolved this LSD1 extension in order to improve the aminoacylation reaction step. The results also showed that the LSD1s from organisms of both groups are dispensable for post-transfer editing. Together, the data provide us with a further understanding of the organization and structure of LeuRS domains.

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A novel pathway for the conversion of homocysteine to methionine in eukaryotes

Biochemical Journal ◽

10.1042/bj3280165 ◽

1997 ◽

Vol 328 (1) ◽

pp. 165-170 ◽

Cited By ~ 6

Author(s):

M. Celia ANTONIO ◽

C. Marta NUNES ◽

Helga REFSUM ◽

K. Abraham ABRAHAM

Keyword(s):

Amino Acid ◽

Crude Extract ◽

Trna Synthetase ◽

Synthetase Activity ◽

Enzyme Complex ◽

Homocysteine Thiolactone ◽

Initiator Trna ◽

Trna Synthetases ◽

Aminoacyl Trna Synthetases ◽

Reticulocyte Lysates

Activation of amino acid homocysteine was compared with that of methionine in rabbit crude liver extracts and purified multi-enzyme complex of aminoacyl-tRNA synthetases. Activation was studied by measuring the incorporation of radioactive amino acid into unlabelled trichloroacetic-acid insoluble materials in the absence of protein synthesis. Homocysteine synthetase activity was found in the crude extract and in the purified multi-enzyme complex of aminoacyl-tRNA synthetases. On a molar basis, the activation of methionine by the crude extract was five times higher than the activation of homocysteine. There was a partial loss of Hcy-tRNA synthetase activity in the purified multi-enzyme complex. Preliminary reconstitution experiments indicated a requirement for an additional factor for Hcy-tRNA synthetase activity. TLC of the amino acid released from tRNA charged with [14C]homocysteine, revealed radioactivity in homocysteine, methionine and homocysteine thiolactone, indicating a conversion of tRNA-attached homocysteine to methionine. Total tRNA was separated on a benzoylated cellulose column into a fraction enriched in initiator tRNA and a methionine-accepting, but initiator tRNA-deficient, fraction. Homocysteine-accepting activity was present only in the initiator tRNA-enriched fraction. Based on the above data we propose that homocysteine activation in reticulocyte lysates, reported previously, also occurs in liver. Activated homocysteine is attached to initiator tRNA and then converted to methionine by a methylating enzyme. In the absence of methylation, tRNA-attached homocysteine is hydrolysed to produce homocysteine thiolactone.

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The Mechanism of Action of Methionyl-tRNA Synthetase from Escherichia coli. Mechanism of the Amino-Acid Activation Reaction Catalyzed by the Native and the Trypsin-Modified Enzymes

European Journal of Biochemistry ◽

10.1111/j.1432-1033.1974.tb03491.x ◽

1974 ◽

Vol 44 (2) ◽

pp. 343-351 ◽

Cited By ~ 63

Author(s):

Sylvain Blanquet ◽

Guy Fayat ◽

Jean-Pierre Waller

Keyword(s):

Escherichia Coli ◽

Amino Acid ◽

Mechanism Of Action ◽

Trna Synthetase ◽

Acid Activation ◽

Activation Reaction ◽

Amino Acid Activation ◽

Methionyl Trna Synthetase

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Evidence that gene G7a in the human major histocompatibility complex encodes valyl-tRNA synthetase

Biochemical Journal ◽

10.1042/bj2780809 ◽

1991 ◽

Vol 278 (3) ◽

pp. 809-816 ◽

Cited By ~ 50

Author(s):

S L Hsieh ◽

R D Campbell

Keyword(s):

Major Histocompatibility Complex ◽

Amino Acid ◽

Molecular Mass ◽

Trna Synthetase ◽

Class I ◽

Major Histocompatibility ◽

High Sequence Identity ◽

Human Major Histocompatibility Complex ◽

Trna Synthetases ◽

Histocompatibility Complex

At least 36 genes have now been located in a 680 kb segment of DNA between the class I and class II multigene families within the class III region of the human major histocompatibility complex on chromosome 6p21.3. The complete nucleotide sequence of the 4.3 kb mRNA of one of these genes, G7a (or BAT6), has been determined from cDNA and genomic clones. The single-copy G7a gene encodes a 1265-amino-acid protein of molecular mass 140,457 Da. Comparison of the derived amino acid sequence of the G7a protein with the National Biomedical Research Foundation protein databases revealed 42% identity in a 250-amino-acid overlap with Bacillus stearothermophilus valyl-tRNA synthetase, 38.0% identity in a 993-amino-acid overlap with Escherichia coli valyl-tRNA synthetase (val RS), and 48.3% identity in a 1043-amino-acid overlap with Saccharomyces cerevisiae valyl-tRNA synthetase. The protein sequence of G7a contains two short consensus sequences, His-Ile-Gly-His and Lys-Met-Ser-Lys-Ser, which is the typical signature structure of class I tRNA synthetases and indicative of the presence of the Rossman fold. In addition, the molecular mass of the G7a protein is the same as that of other mammalian valyl-tRNA synthetases. These features and the high sequence identity with yeast valyl-tRNA synthetase strongly support the fact that the G7a gene, located within the major histocompatibility complex, encodes the human valyl-tRNA synthetase.

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