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KMID : 0364820130490020105
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Korean Journal of Microbiology
2013 Volume.49 No. 2 p.105 ~ p.111
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Mutational Analysis Elucidates the Role of Conserved 237 Arginine in 23S rRNA Methylation, Which is in the Concave Cleft Region of ErmSF
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Jin Hyung-Jong
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Abstract
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The Erm family of adenine-N6methyltransferases (MTases) is responsible for the development of resistance to
macrolide-lincosamide-streptogramin B antibiotics through the methylation of 23S ribosomal RNA. Recently, it has
been proposed that well conserved amino acids in ErnC’ located in concave cleft between N-terminal ‘catalytic’
domain and C-terminal ‘RNA-binding’ domain interacts with substrate RNA. We carried out the site-directed mutagenesis and studied the function of the ErmSF R237 mutant in vitro and in vivo. R237 amino acid residue is located in the concave cleft between two domains. Furthermore this residue is very highly conserved in almost all the Erm family. Purified mutant protein exhibited only 51% enzyme activity compared to wild-type. Escherichia coli with R237A mutant protein compared to the wild-type protein expressing E. coli did not show any difference in its MIC(minimal inhibitory concentration) suggesting that even with lowered enzyme activity, mutant protein was able to efficiently methylate 23S rRNA to confer the resistance on E. coli expressing this protein. But this observation strongly suggests that R237 of ErmSF probably interacts with substrate RNA affecting enzyme activity significantly
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KEYWORD
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antibiotic resistance factor protein, in vivo, in vitro activity test, MLSb (macrolide-lincosamidestreptogramin B) site-direc
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