Detection of physical interactions by immunoprecipitation of FLAG- and HA-tagged proteins expressed at the his-3 locus in Neurospora crassa

Abstract

Protein function is often regulated through interactions with other protein(s) or by post-translational modifications. To understand these mechanisms, it is useful to utilize antibodies. However, it is not always certain whether a good antibody can be made for this purpose. The use of epitope tags eliminates the troubles associated with raising antibodies. In this report, we present a method to detect interactions between proteins by using two types of epitope-tagged proteins, FLAGand HA-tagged proteins in Neurospora. These constructs were introduced at and expressed from the his-3 locus in different strains. To examine protein-protein interactions, heterokaryons between these strains were constructed. We conclude that this strategy is a useful tool to investigate protein function and protein interactions.