A generic protocol for the affinity-purification of native macromolecular complexes from poxvirus-infected cells

Abstract

Summary: The functional and structural characterization of macromolecular complexes requires protocols for their native isolation. Here, we describe a protocol for this task based on the recombinant poxvirus Vaccinia expressing tagged proteins of interest in infected cells. Tagged proteins and their interactors can then be isolated via affinity chromatography. The procedure is illustrated for the Vaccinia virus encoded multi-subunit RNA polymerase. Our protocol also allows the expression and isolation of heterologous proteins and hence is suitable for a broader application.For complete details on the use and execution of this profile, please refer to Grimm et al. (2019).

Keywords

• Cell Biology
• Microbiology
• Molecular Biology
• Protein Biochemistry
• Protein expression and purification