Distinction between the Cfr methyltransferase conferring antibiotic resistance and the housekeeping RlmN
Gemma C Atkinson1, Lykke H Hansen, Tanel Tenson
1Institute of Technology, University of Tartu, Tartu, Estonia.
Abstract:
The cfr gene encodes the Cfr methyltransferase that primarily methylates C-8 in A2503 of 23S rRNA in the peptidyl transferase region of bacterial ribosomes. The methylation provides resistance to six classes of antibiotics of clinical and veterinary importance. The rlmN gene encodes the RlmN methyltransferase that methylates C-2 in A2503 in 23S rRNA and A37 in tRNA, but RlmN does not significantly influence antibiotic resistance. The enzymes are homologous and use the same mechanism involving radical S-adenosyl methionine to methylate RNA via an intermediate involving a methylated cysteine in the enzyme and a transient cross-linking to the RNA, but they differ in which carbon atom in the adenine they methylate. Comparative sequence analysis identifies differentially conserved residues that indicate functional sequence divergence between the two classes of Cfr- and RlmN-like sequences. The differentiation between the two classes is supported by previous and new experimental evidence from antibiotic resistance, primer extensions, and mass spectrometry. Finally, evolutionary aspects of the distribution of Cfr- and RlmN-like enzymes are discussed.
Insights
The Cfr methyltransferase confers antibiotic resistance by modifying 23S rRNA. Comparative analysis reveals functional divergence between Cfr and RlmN enzymes, impacting their roles in bacterial ribosomes and antibiotic resistance.
Area of Science:
- Microbiology
- Molecular Biology
- Biochemistry
Background:
- The cfr gene encodes Cfr methyltransferase, which methylates 23S rRNA at C-8, conferring resistance to multiple antibiotic classes.
- The homologous rlmN gene encodes RlmN methyltransferase, methylating 23S rRNA at C-2 and tRNA at A37, with minimal impact on antibiotic resistance.
Purpose of the Study:
- To investigate the functional divergence between Cfr and RlmN methyltransferases.
- To identify sequence variations correlating with functional differences in antibiotic resistance mechanisms.
Main Methods:
- Comparative sequence analysis of Cfr and RlmN enzymes.
- Experimental validation using antibiotic resistance assays, primer extensions, and mass spectrometry.
Main Results:
- Differential conservation of residues between Cfr and RlmN sequences suggests functional divergence.
- Experimental data supports distinct roles in antibiotic resistance and methylation sites.
Conclusions:
- Cfr and RlmN enzymes, despite homology, exhibit functional divergence in RNA methylation, impacting antibiotic resistance.
- Sequence variations provide insights into the evolution and distribution of these methyltransferases.
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