Abstract
Our laboratory previously identified a cDNA clone encoding the small heat shock protein HSPB3. A hybridoma that produces monoclonal antibody specific to HSPB3 was also generated. HSPB3 showed an extremely high expression level by western blot analysis in rat heart tissue, and could be detected in a less extent in skeletal muscle, brain and kidney. To further understand the physiological function of HSPB3, we screened the human heart cDNA library with the yeast two-hybrid system and showed that actin, which is expressed at a high level and plays an important role in muscle tissue, can interact with HSPB3. To identify the protein interaction domain, we made a series of truncated mutation of these two proteins and assayed their interaction by using the yeast two-hybrid system. The results indicated that the region containing amino acids 24-48 of HSPB3 is critical for interaction with actin. In actin, the C-terminal portion, especially amino acids 302-337, is necessary for its interaction with HSPB3. Nevertheless, site-specific mutagenesis of the amino acids 324-327, which involved in actin polymerization did not affect the interaction. Therefore, the exact region required for the HSPB3-actin interaction remains to be identified. In conclusion, our results have provided basic information required for a deeper insight into the properties and the functional role of the small heat shock protein HSPB3.