PMID:8065908
| Citation |
Borel, F, Vincent, C, Leberman, R and Härtlein, M (1994) Seryl-tRNA synthetase from Escherichia coli: implication of its N-terminal domain in aminoacylation activity and specificity. Nucleic Acids Res. 22:2963-9 |
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| Abstract |
Escherichia coli seryl-tRNA synthetase (SerRS) a dimeric class II aminoacyl-tRNA synthetase with two structural domains charges specifically the five iso-acceptor tRNA(ser) as well as the tRNA(sec) (selC product) of E. coli. The N-terminal domain is a 60 A long arm-like coiled coil structure built of 2 long antiparallel a-h helices, whereas the C-terminal domain is a alpha-beta structure. A deletion of the N-terminal arm of the enzyme does not affect the amino acid activation step of the reaction, but reduces dramatically amino-acylation activity. The Kcat/Km value for the mutant enzyme is reduced by more than 4 orders of magnitude, with a nearly 30 fold increased Km value for tRNA(ser). An only slightly truncated mutant form (16 amino acids of the tip of the arm replaced by a glycine) has an intermediate aminoacylation activity. Both mutant synthetases have lost their specificity for tRNA(ser) and charge also non-cognate type 1 tRNA(s). Our results support the hypothesis that class II synthetases have evolved from an ancestral catalytic core enzyme by adding non-catalytic N-terminal or C-terminal tRNA binding (specificity) domains which act as determinants for cognate and anti-determinants for non-cognate tRNAs. |
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| Keywords |
Acylation; Adenosine Triphosphate/metabolism; Binding Sites; Escherichia coli/enzymology; Gene Deletion; Kinetics; Macromolecular Substances; Models, Molecular; Mutagenesis, Site-Directed; Protein Structure, Secondary; RNA, Transfer, Ser/metabolism; Serine/metabolism; Serine-tRNA Ligase/chemistry; Serine-tRNA Ligase/genetics; Serine-tRNA Ligase/metabolism; Structure-Activity Relationship |
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