Protocol for purification, crystallization, and structure determination of the mpox virus mRNA cap N7 methyltransferase complex
Summary: The mpox virus (MPXV) mRNA cap N7 methyltransferase (RNMT) methylates guanosine at mRNA 5′-cap N7 positions to enable immune evasion. Here, we present a protocol for E1CTD-E12 complex purification and crystallization. We describe steps for rational sequence design of the complex, co-express...
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| Main Authors: | , , , |
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| Format: | Article |
| Language: | English |
| Published: |
Elsevier
2025-06-01
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| Series: | STAR Protocols |
| Subjects: | |
| Online Access: | http://www.sciencedirect.com/science/article/pii/S2666166725002515 |
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| Summary: | Summary: The mpox virus (MPXV) mRNA cap N7 methyltransferase (RNMT) methylates guanosine at mRNA 5′-cap N7 positions to enable immune evasion. Here, we present a protocol for E1CTD-E12 complex purification and crystallization. We describe steps for rational sequence design of the complex, co-expression in E. coli, affinity chromatography purification, gel filtration, and crystallization optimization using vapor diffusion. We further outline X-ray diffraction data collection and structure determination. This reproducible framework enables structural analysis of viral mRNA-modifying enzyme complexes.For complete details on the use and execution of this protocol, please refer to Chen et al.1 : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics. |
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| ISSN: | 2666-1667 |