Characterization of alpha-crystallin-plasma membrane binding

alpha-Crystallin, a large lenticular protein complex made up of two related subunits (alpha A- and alpha B-crystallin), is known to associate increasi ngly with fiber cell plasma membranes with age and/or the onset of cataract , To understand better the binding mechanism, we developed a sensitive memb rane binding assay using lens plasma membranes and recombinant human alpha A- and alpha B-crystallins conjugated to a small fluorescent tag (Alexa350( R)). Both alpha A and alpha B homopolymer complexes, as well as a reconstit uted 3:1 heteromeric complex, bind to lens membranes in a specific, saturab le, and partially irreversible manner that is sensitive to both time and te mperature. The amount of alpha-crystallin that binds to the membrane increa ses under acidic pH conditions and upon removal of exposed intrinsic membra ne protein domains but is not affected at high ionic strength, suggesting t hat alpha-crystallin binds to the fiber cell plasma membranes mainly throug h hydrophobic interactions. The binding capacity and affinity for the recon stituted 3:1 heteromeric complex were measured to be 3.45 +/- 0.11 ng/mu g of membrane and 4.57 +/- 0.50 x 10(-4) mu g(-1) of membrane, respectively, The present membrane binding data support the hypothesis that the physical properties of a mixed alpha-crystallin complex may hold particular relevanc e for the function of alpha-crystallin within the lens.