Crystal structure and functional analysis of mycobacterial erythromycin resistance methyltransferase Erm38 reveals its RNA-binding site
Erythromycin resistance methyltransferases (Erms) confer resistance to macrolide, lincosamide, and streptogramin antibiotics in Gram-positive bacteria and mycobacteria. Although structural information for ErmAM, ErmC, and ErmE exists from Gram-positive bacteria, little is known about the Erms in myc...
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| Format: | Journal Article |
| Language: | English |
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2022
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| Online Access: | https://hdl.handle.net/10356/161111 |
| _version_ | 1826122588468805632 |
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| author | Goh, Boon Chong Xiang, Xinyu Lescar, Julien Dedon, Peter C. |
| author2 | School of Biological Sciences |
| author_facet | School of Biological Sciences Goh, Boon Chong Xiang, Xinyu Lescar, Julien Dedon, Peter C. |
| author_sort | Goh, Boon Chong |
| collection | NTU |
| description | Erythromycin resistance methyltransferases (Erms) confer resistance to macrolide, lincosamide, and streptogramin antibiotics in Gram-positive bacteria and mycobacteria. Although structural information for ErmAM, ErmC, and ErmE exists from Gram-positive bacteria, little is known about the Erms in mycobacteria, as there are limited biochemical data and no structures available. Here, we present crystal structures of Erm38 from Mycobacterium smegmatis in apoprotein and cofactor-bound forms. Based on structural analysis and mutagenesis, we identified several catalytically critical, positively charged residues at a putative RNA-binding site. We found that mutation of any of these sites is sufficient to abolish methylation activity, whereas the corresponding RNA-binding affinity of Erm38 remains unchanged. The methylation reaction thus appears to require a precise ensemble of amino acids to accurately position the RNA substrate, such that the target nucleotide can be methylated. In addition, we computationally constructed a model of Erm38 in complex with a 32-mer RNA substrate. This model shows the RNA substrate stably bound to Erm38 by a patch of positively charged residues. Furthermore, a π-π stacking interaction between a key aromatic residue of Erm38 and a target adenine of the RNA substrate forms a critical interaction needed for methylation. Taken together, these data provide valuable insights into Erm-RNA interactions, which will aid subsequent structure-based drug design efforts. |
| first_indexed | 2024-10-01T05:50:36Z |
| format | Journal Article |
| id | ntu-10356/161111 |
| institution | Nanyang Technological University |
| language | English |
| last_indexed | 2024-10-01T05:50:36Z |
| publishDate | 2022 |
| record_format | dspace |
| spelling | ntu-10356/1611112023-02-28T17:13:28Z Crystal structure and functional analysis of mycobacterial erythromycin resistance methyltransferase Erm38 reveals its RNA-binding site Goh, Boon Chong Xiang, Xinyu Lescar, Julien Dedon, Peter C. School of Biological Sciences NTU Institute of Structural Biology Science::Biological sciences Antibiotic Resistance RNA-Binding Protein Erythromycin resistance methyltransferases (Erms) confer resistance to macrolide, lincosamide, and streptogramin antibiotics in Gram-positive bacteria and mycobacteria. Although structural information for ErmAM, ErmC, and ErmE exists from Gram-positive bacteria, little is known about the Erms in mycobacteria, as there are limited biochemical data and no structures available. Here, we present crystal structures of Erm38 from Mycobacterium smegmatis in apoprotein and cofactor-bound forms. Based on structural analysis and mutagenesis, we identified several catalytically critical, positively charged residues at a putative RNA-binding site. We found that mutation of any of these sites is sufficient to abolish methylation activity, whereas the corresponding RNA-binding affinity of Erm38 remains unchanged. The methylation reaction thus appears to require a precise ensemble of amino acids to accurately position the RNA substrate, such that the target nucleotide can be methylated. In addition, we computationally constructed a model of Erm38 in complex with a 32-mer RNA substrate. This model shows the RNA substrate stably bound to Erm38 by a patch of positively charged residues. Furthermore, a π-π stacking interaction between a key aromatic residue of Erm38 and a target adenine of the RNA substrate forms a critical interaction needed for methylation. Taken together, these data provide valuable insights into Erm-RNA interactions, which will aid subsequent structure-based drug design efforts. Agency for Science, Technology and Research (A*STAR) National Research Foundation (NRF) Singapore-MIT Alliance for Research and Technology (SMART) Published version This work was supported by grants from the National Research Foundation of Singapore through the Singapore-MIT Alliance for Research and Technology (SMART) Antimicrobial Resistance Interdisciplinary Research Group and the SMART Innovation Centre (ING-000772 BIO IGN), and by a grant from the A*STAR BMRC Therapeutics Development Review (H19H0G1003/TOR 005–018). 2022-08-16T04:37:02Z 2022-08-16T04:37:02Z 2022 Journal Article Goh, B. C., Xiang, X., Lescar, J. & Dedon, P. C. (2022). Crystal structure and functional analysis of mycobacterial erythromycin resistance methyltransferase Erm38 reveals its RNA-binding site. Journal of Biological Chemistry, 298(2), 101571-. https://dx.doi.org/10.1016/j.jbc.2022.101571 0021-9258 https://hdl.handle.net/10356/161111 10.1016/j.jbc.2022.101571 35007529 2-s2.0-85125006923 2 298 101571 en ING-000772 BIO IGN H19H0G1003/TOR 005–018 Journal of Biological Chemistry © 2022 The Authors. Published by Elsevier Inc on behalf of American Society for Biochemistry and Molecular Biology. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/). application/pdf |
| spellingShingle | Science::Biological sciences Antibiotic Resistance RNA-Binding Protein Goh, Boon Chong Xiang, Xinyu Lescar, Julien Dedon, Peter C. Crystal structure and functional analysis of mycobacterial erythromycin resistance methyltransferase Erm38 reveals its RNA-binding site |
| title | Crystal structure and functional analysis of mycobacterial erythromycin resistance methyltransferase Erm38 reveals its RNA-binding site |
| title_full | Crystal structure and functional analysis of mycobacterial erythromycin resistance methyltransferase Erm38 reveals its RNA-binding site |
| title_fullStr | Crystal structure and functional analysis of mycobacterial erythromycin resistance methyltransferase Erm38 reveals its RNA-binding site |
| title_full_unstemmed | Crystal structure and functional analysis of mycobacterial erythromycin resistance methyltransferase Erm38 reveals its RNA-binding site |
| title_short | Crystal structure and functional analysis of mycobacterial erythromycin resistance methyltransferase Erm38 reveals its RNA-binding site |
| title_sort | crystal structure and functional analysis of mycobacterial erythromycin resistance methyltransferase erm38 reveals its rna binding site |
| topic | Science::Biological sciences Antibiotic Resistance RNA-Binding Protein |
| url | https://hdl.handle.net/10356/161111 |
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