Structure -function relationship of arginyl-tRNA synthetase from Escherichia coli: isolation and characterization of the argS mutation MA5002 - CNRS - Centre national de la recherche scientifique Accéder directement au contenu
Article Dans Une Revue Nucleic Acids Research Année : 1990

Structure -function relationship of arginyl-tRNA synthetase from Escherichia coli: isolation and characterization of the argS mutation MA5002

Gilbert Eriani
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Guy Dirheimer
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Jean Gangloff
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Résumé

The Escherichia coli Kl 2 argS MA5002 mutant appears to have a functionally altered arginyl-tRNA synthetase (ArgRS). The gene coding for this enzyme was isolated from E.coli genomic DNA using the PCR procedure and inserted into a pUC18 multicopy vector. Sequencing revealed that it differs from the wildtype ArgRS structural gene only by one mutation: a replacement of a C by an A residue which results in substitution of an arginine by a serine at position 134, located two residues downstream from the HVGH consensus sequence. As compared to the genomic enzyme level, this recombinant vector, containing the mutated gene, produces in E.coli JM103, about 100 times as much modified ArgRS. This enzyme was obtained nearly pure after only two chromatographic steps; it exhibits a 4-6 times as low activity and a 5 times as high Km value for ATP as the wildtype enzyme in the aminoacylation and ATP-PPi reactions; Km values for arginine and tRNAA'g remained unaltered. The position of this mutation and its effect on enzymatic properties suggest the implication of arginine 134 in ATP binding as well as in the activation catalytic process.
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hal-02465276 , version 1 (03-02-2020)

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  • HAL Id : hal-02465276 , version 1

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Gilbert Eriani, Guy Dirheimer, Jean Gangloff. Structure -function relationship of arginyl-tRNA synthetase from Escherichia coli: isolation and characterization of the argS mutation MA5002. Nucleic Acids Research, 1990. ⟨hal-02465276⟩

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