Characterization of dimer subunits of intermediate filament proteins.
The fundamental subunit of the various types of intermediate-sized filaments (IF) has been shown to be a tetramer that is thought to represent a double dimer, i.e. an array of two laterally packed coiled-coils of alpha-helices. The two-chain state of intact IF proteins had up to this point not been isolated and characterized as has been done for other fibrous alpha-helical coiled-coil proteins. Using buffers containing 3 M-guanidinium hydrochloride we prepared dimers by depolymerization of IF or by reconstitution from fully denatured molecules. Dimers of desmin (from chicken gizzard), vimentin (from bovine lens tissue and cultured human fibroblasts) and the neurofilament protein NF-L (from bovine brain) as well as in vitro formed homodimers of human and rat cytokeratins numbers 8 (A), 18 (D) and 19 ("40K"), are characterized by ultracentrifugation techniques (sedimentation velocity and equilibrium), electron microscopy and chemical cross-linking. The results show that IF proteins from discrete complexes of two polypeptide chains in parallel orientation and probably in coiled-coil configuration, which apparently have a high tendency to further associate into double dimers. Implications of these results for concepts of IF organization and IF protein assembly are discussed.
D000818:Animals; D002645:Chickens; D002851:Chromatography, High Pressure Liquid; D002942:Circular Dichroism; D003432:Cross-Linking Reagents; D003893:Desmin; D007381:Intermediate Filament Proteins; D007633:Keratins; D046911:Macromolecular Substances; D008854:Microscopy, Electron; D016900:Neurofilament Proteins; D014461:Ultracentrifugation; D014746:Vimentin
D003432:Cross-Linking Reagents; D003893:Desmin; D007381:Intermediate Filament Proteins; D046911:Macromolecular Substances; D016900:Neurofilament Proteins; D014746:Vimentin; C044882:neurofilament protein L; D007633:Keratins