Monitoring and purification of proteins using bovine papillomavirus E2 epitope tags

We describe here the use of two newly mapped bovine papillomavirus type I (BPV-I) E2 protein epitopes as rags. We constucted several vector plasmids for overexpression as Ic ell as for moderate expression of single- or double-tugged proteins in either Escherichia coli or eukaryotic cells. The new tags were fused to several proteins, and the activity of the tugged proteins was tested in different assays. The rags were shown nor to interfere, with the function of these proteins in I vivo and in vitro. Interaction of the monoclonal antibodies 3F12 and 1E2 with their respective epitopes M as specific and had high affinity in a variety of conditions. We have demonstrated that the 3F12 antibody-epitope interaction tolerates high salt concentrations rip to 2 M. This permits immunoprecipitation and immunopurification of the tagged proteins in high-salt buffers and reduction of the nonspecific binding of the contaminating proteins. We also provide a protocol for DNA binding and DNase I footprinting assays using the tagged resin-bound DNA-binding proteins. The BPV-I E2-derived tags can be recommended as useful tools for detection and purification of proteins.