Cell-specific domains of glial- and muscle-type intermediate filament proteins. Immunoaffinity chromatography and immunoblotting study of GFA protein and desmin.
In order to localize the cell specific domains of glial- and muscle-type intermediate filaments, the purified subunits (bovine GFA protein and chicken desmin) were fragmented and the digests passed through immunoaffinity columns or stained by the immunoblotting procedure to determine which fragments reacted with the monospecific polyvalent antisera. The following fragments were found immunoreactive according to these criteria: 30 K (GFA) and 33 K (desmin) N-bromosuccinimide fragments (tryptophan cleavage); 35 K (GFA) and 39 K (desmin) 2-nitro-5-thiocyanobenzoic acid fragments (cysteine cleavage); 18 K (GFA) and 9 K (desmin) cyanogen bromide fragments. Fragmentation of GFA protein was also accomplished using proteolytic digestion with chymotrypsin and trypsin. Two resistant core polypeptides, one about 37 K and stable in the chymotryptic digests and one about 21 K and stable in the tryptic digests bound specifically to the immunoaffinity columns. The 21 K tryptic fragment was found to contain the 18 K cyanogen bromide fragment. The fragmentation patterns support recently published structural domain models for intermediate filament proteins. The immunochemical findings indicate that the immunoreactive regions of GFA protein are located in the aminoterminal region of the middle domain of these models (coil I), while they appear to be situated in the aminoterminal headpiece of the protein in the case of desmin.
D000595:Amino Acid Sequence; D000818:Animals; D002417:Cattle; D002645:Chickens; D002846:Chromatography, Affinity; D003488:Cyanogen Bromide; D003893:Desmin / Q000032:analysis*; D005904:Glial Fibrillary Acidic Protein / Q000032:analysis*; D006868:Hydrolysis; D007120:Immunochemistry; D008958:Models, Molecular; D009132:Muscles / Q000032:analysis; D010446:Peptide Fragments / Q000032:analysis
D003893:Desmin; D005904:Glial Fibrillary Acidic Protein; D010446:Peptide Fragments; D003488:Cyanogen Bromide