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A structural basis for the antibiotic resistance conferred by an N1-methylation of A1408 in 16S rRNA
Hiroki Kanazawa1, Fumika Baba1, Mai Koganei1
1Graduate School of Science and Technology, Sophia University, 7-1 Kioi-cho, Chiyoda-ku, 102-8554 Tokyo, Japan.
Abstract:
The aminoglycoside resistance conferred by an N1-methylation of A1408 in 16S rRNA by a novel plasmid-mediated methyltransferase NpmA can be a future health threat. In the present study, we have determined crystal structures of the bacterial ribosomal decoding A site with an A1408m1A antibiotic-resistance mutation both in the presence and absence of aminoglycosides. G418 and paromomycin both possessing a 6'-OH group specifically bind to the mutant A site and disturb its function as a molecular switch in the decoding process. On the other hand, binding of gentamicin with a 6'-NH3+ group to the mutant A site could not be observed in the present crystal structure. These observations agree with the minimum inhibitory concentration of aminoglycosides against Escherichia coli. In addition, one of our crystal structures suggests a possible conformational change of A1408 during the N1-methylation reaction by NpmA. The structural information obtained explains how bacteria acquire resistance against aminoglycosides along with a minimum of fitness cost by the N1-methylation of A1408 and provides novel information for designing the next-generation aminoglycoside.
Insights
A novel enzyme, NpmA, causes aminoglycoside resistance by modifying bacterial 16S rRNA. Structural studies reveal how specific antibiotics bind to the resistant mutation, aiding future drug design.
Area of Science:
- Molecular Biology
- Structural Biology
- Antimicrobial Resistance
Background:
- Aminoglycoside antibiotics are crucial for treating bacterial infections.
- Emerging resistance mechanisms, like N1-methylation of A1408 in 16S rRNA by NpmA, pose a significant public health threat.
- Understanding the structural basis of this resistance is vital for developing new therapeutic strategies.
Purpose of the Study:
- To determine the crystal structures of the bacterial ribosomal decoding A site with the A1408m1A mutation in the presence and absence of aminoglycosides.
- To elucidate the molecular interactions between specific aminoglycosides and the resistant ribosomal site.
- To provide structural insights into the mechanism of NpmA-mediated aminoglycoside resistance.
Main Methods:
- X-ray crystallography to obtain high-resolution structures of the ribosomal A site.
- Biochemical assays to assess aminoglycoside binding and functional impact.
- Minimum inhibitory concentration (MIC) studies to correlate structural findings with antibiotic efficacy.
Main Results:
- Crystal structures revealed specific binding of G418 and paromomycin (with a 6'-OH group) to the A1408m1A mutant A site, disrupting its function.
- Gentamicin (with a 6'-NH3+ group) did not show binding to the mutant A site in the crystal structure.
- Structural data supports the low fitness cost associated with NpmA-mediated resistance and suggests a conformational change during methylation.
Conclusions:
- The N1-methylation of A1408 by NpmA confers resistance by altering aminoglycoside binding interactions within the ribosomal A site.
- Structural insights explain the differential efficacy of various aminoglycosides against resistant strains.
- This study provides a foundation for designing next-generation aminoglycosides effective against NpmA-mediated resistance.