Purification of recombinant proteins and study of protein interaction by epitope tagging

A protein molecule can be engineered to include a short stretch of residues corresponding to an epitope to facilitate its subsequent biochemical and immunological analysis; a technique often referred to as "epitope tagging." This unit presents a protocol for small-scale immunoprecipitation of epitope-tagged recombinant proteins expressed in transiently transfected mammalian cells. The immunoprecipitant can then be analyzed by SDS-PAGE. An immunoprecipitation protocol is also provided that has been optimized for use with a baculovirus overexpression system. An Alternate Protocol describes how multisubunit complexes can be assembled by starting with a core protein affixed to beads via an epitope tag, and adding the other members of the complex in a stepwise manner.