Alpha-crystallin, a major eye lens protein of vertebrates has been characterized as a molecular chaperone based on its ability to inhibit the aggregation of proteins undergoing thermal denaturation (Horwitz, J., Proc. Natl. Acad. Sci. USA 1992, 89, 10449-10453). To understand the mechanisms underlying this chaperone-like activity, the present study addressed molecular interactions between alpha-crystallin and its target proteins. Using carbonic anhydrase as a model target protein, we demonstrate complex formation between the 2 proteins upon heating, as assessed by the criteria of agarose gel electrophoresis, immunoprecipitation, ultrafiltration and gel filtration chromatography. The complex of alpha-crystallin and carbonic anhydrase is stable, at room temperature and at 4 degrees C, for over 18 hours, and is non-covalent in nature. The results also indicate that alpha-crystallin binds the early non-native form of the target protein.