Isolation and characterization of keratin-like proteins from cultured cells with fibroblastic morphology.

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Abstract
Intermediate filaments (IF) isolated from a variety of cultured cells, conventionally described as fibrolasts, are composed predominately of proteins of MW of 54,000 and/or 55,000. Less than 15% of the protein found in native IF preparations from these cells is composed of 3-4 polypeptides of MW 60,000-70,000. Some biochemical and immunological properties of these proteins isolated from BHK-21 and mouse 3T3 cells were investigated. They are capable of forming paracrystals that exhibit a light/dark banding pattern when negatively stained with uranyl acetate. The dark bands are composed of longitudinally aligned .apprx. 2-nm-diameter filaments. The center-to-center spacing between either dark or light bands is 37-40 nm. These dimensions are consistent with the secondary structure of IF polypeptides and suggest that the dark bands represent lateral alignment of .alpha.-helical coiled-coil domains. Immunoblotting, secondary structure, as well as amino acid composition data indicate that the 60,000-70,000-MW paracrystal polypeptides are similar to keratin. Polypeptides with biochemical and immunological properties of epidermal keratin are present in cells normally considered to be fibroblasts.