Work

Protocol for purification, crystallization, and structure determination of the mpox virus mRNA cap N7 methyltransferase complex

Year: 2025

Type: article

Abstract: 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 E1 CTD -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

Source: STAR Protocols

DOI: 10.1016/j.xpro.2025.103845

Authors: Qingqing Guo, Anke Chen, Jin Ding, Jixi Li

Institutions: Fudan University, Pudong Medical Center, Second Military Medical University

Language: English

Topic: Enzyme Structure and Function

Subfield: Materials Chemistry

Field: Materials Science

Domain: Physical Sciences

Open Access status: gold

Estimated APC paid: $1,550

Funder: National Natural Science Foundation of China