Identification of noncollagenous components of calf lens capsule: evaluation of their adhesion-promoting activity.

Extraction of calf anterior and posterior lens capsules with 5 M guanidine HCI resulted in the solubilization of protein (12% of total) with a noncollagenous amino acid composition leaving behind the collagen matrix. Polyacrylamide gel electrophoresis of the solubilized material revealed a number of components, all of which were susceptible to trypsin but resistant to collagenase digestion. Fractionation of the extracted proteins by Sepharose CL-6B filtration as well as by affinity chromatography was undertaken, and laminin, fibronectin, entactin, and beta-crystallin were identified by electrophoresis and solid-phase radioimmunoassays in both anterior and posterior capsules. An entactin (Mr = 150,000), which constituted the most prominent component on electrophoresis, was purified after Sepharose CL-6B filtration by a two-step lectin affinity chromatography procedure, which was based on the failure of this protein to bind to Bandeiraea simplicifolia I but its positive reactivity with wheat germ lectin. Neither the mixture of proteins extracted from lens capsules by guanidine nor fractions prepared therefrom were able to enhance lens epithelial cell attachment to type I or type IV collagen-coated surfaces or to guanidine-prepared lens capsules; adhesion-stimulating activity could not be demonstrated even when cycloheximide-treated cells were employed. Furthermore, the cells were observed to attach as effectively to guanidine-extracted as to native capsules. These observations indicate that noncollagenous proteins are not essential for the in vitro attachment of epithelial cells to lens capsule; it appears that the collagen component itself provides an optimal surface for cell-basement membrane interaction.