Epidermal alpha-keratin is neutral-buffer-soluble and forms intermediate filaments under physiological conditions in vitro.
Undenatured bovine epidermal alpha-keratin has been solubilized in a low-ionic-strength buffer at physiological pH (5 mM Tris-HCl/25 mM 2-mercaptoethanol (pH 7.5). The particles in this buffer were multimeric, retaining their characteristic polypeptide chain composition and alpha-helical coiled-coil structure. They were shown by sucrose density gradient centrifugation to be in true solution and to have a narrow size distribution. Upon the addition to this solution of monovalent or divalent cations up to physiological concentrations, the alpha-keratin rapidly assembled into intermediate filaments which showed a high tendency to aggregate laterally and disassembled if returned to low-ionic-strength conditions. This behaviour closely resembles that of other intermediate filament proteins, but it is the first time that alpha-keratins have been shown to be neutral-buffer-soluble and to assemble from such solutions into intermediate filaments under physiological conditions in vitro. This is in direct contrast to the reported properties of alpha-keratins after urea denaturation and the system appears to be appropriate for studying aspects of alpha-keratin intermediate filament formation.
D000818:Animals; D002021:Buffers; D002412:Cations; D002417:Cattle; D002499:Centrifugation, Density Gradient; D003599:Cytoskeleton / Q000378:metabolism*; D004817:Epidermis / Q000032:analysis*; D006863:Hydrogen-Ion Concentration; D007382:Intermediate Filaments / Q000378:metabolism*; D007633:Keratins / Q000378:metabolism*; D046911:Macromolecular Substances; D008854:Microscopy, Electron; D009994:Osmolar Concentration; D011487:Protein Conformation; D012995:Solubility
D002021:Buffers; D002412:Cations; D046911:Macromolecular Substances; D007633:Keratins