Protein purification, crystallization, and structure determination of human DEAD-box RNA helicase DDX21 in different unwinding states

Abstract

Summary: RNA helicase DDX21 plays vital roles in ribosomal RNA processing and the regulation of host innate immunity during virus infection. Here, we describe the optimized protocols for nucleic acid-free protein purification and crystallization of DDX21 in its different unwinding states. Rational design of the flexible region within the helicase core, and biophysical approach to characterize interactions between DDX21 and RNA, leads to successful crystallization of DDX21. This protocol can be applied to the crystallography of other DExD/H-box RNA helicases.For complete details on the use and execution of this protocol, please refer to Chen et al. (2020). : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.

Keywords

• Molecular Biology
• Protein Biochemistry
• Protein expression and purification
• Structural Biology
• X-ray Crystallography