Amino acid activation analysis of primitive aminoacyl-tRNA synthetases encoded by both strands of a single gene using the malachite green assay
| Autor: | Yu Sugita, Tomoko Shoji, Hiromi Mutsuro-Aoki, Riku Otsuka, Narumi Yamaki, Nana Suganuma, Ayaka Minobe, Takuya Umehara, Koji Tamura, Haruka Takano, Kazaha Onodera |
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| Rok vydání: | 2021 |
| Předmět: |
Statistics and Probability
Alanine chemistry.chemical_classification Amino acid activation Aminoacyl tRNA synthetase Applied Mathematics General Medicine Pyrophosphate Protein Structure Secondary General Biochemistry Genetics and Molecular Biology Amino acid Amino Acyl-tRNA Synthetases chemistry.chemical_compound Bacterial Proteins chemistry Biochemistry Modeling and Simulation Rosaniline Dyes TRNA aminoacylation Amino Acid Sequence Amino Acids Binding site Histidine |
| Zdroj: | Biosystems. 208:104481 |
| ISSN: | 0303-2647 |
| DOI: | 10.1016/j.biosystems.2021.104481 |
| Popis: | The Rodin-Ohno hypothesis postulates that two classes of aminoacyl-tRNA synthetases were encoded complementary to double-stranded DNA. Particularly, Geobacillus stearothermophilus tryptophanyl-tRNA synthetase (TrpRS, belonging to class I) and Escherichia coli histidyl-tRNA synthetase (HisRS, belonging to class II) show high complementarity of the middle base of the codons in the mRNA sequence encoding each ATP binding site. Here, for the reported 46-residue peptides designed from the three-dimensional structures of TrpRS and HisRS, amino acid activation analysis was performed using the malachite green assay, which detects the pyrophosphate departing from ATP in the forward reaction of the first step of tRNA aminoacylation. A maltose-binding protein fusion with the 46 residues of TrpRS (TrpRS46mer) exhibited high activation capacity for several amino acids in the presence of ATP and amino acids, but the activity of an alanine substitution mutant of the first histidine in the HIGH motif (TrpRS46merH15A) was largely reduced. In contrast, pyrophosphate release by HisRS46mer in the histidine activation step was lower than that in the case of TrpRS46mer. Both HisRS46mer and the alanine mutant at the 113th arginine (HisRS46merR113A) showed slightly higher levels of pyrophosphate release than the maltose-binding protein alone. These results do not rule out the Rodin-Ohno hypothesis, but may suggest the necessity of establishing unique evolutionary models from different perspectives. |
| Databáze: | OpenAIRE |
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