Purification of a 190 kDa protein from smooth muscle: relationship to talin.
Several studies of vinculin-binding proteins have described a 190 kDa protein in chicken gizzard smooth muscle which binds radioiodinated vinculin. We have purified and studied the 190 kDa protein from chicken gizzard smooth muscle. By indirect immunofluorescence, an antiserum raised against the 190 kDa protein stains adhesion plaques (focal contacts), ruffling membranes, and fibrillar streaks on the dorsal and ventral surfaces of fibroblasts. Both the binding to vinculin and the location of the protein in fibroblasts are properties shared with talin, a 215 kDa protein in smooth muscle and fibroblasts. Because antisera against talin and the 190 kDa cross-react the relationship of these two proteins has been investigated further. Upon prolonged storage at 4 degrees C, purified talin degrades into a 190 kDa fragment. A 190 kDa fragment is also generated from talin by the Staphylococcus aureus V-8 proteinase and by trypsin. Comparison of partial peptide maps of talin and the 190 kDa protein reveal that the proteins are very similar and when the 190 kDa fragment of talin is compared with the purified 190 kDa protein by partial proteolytic digestion no differences are found in the pattern of peptides generated. In addition, the amount of 190 kDa protein detected in muscle tissues excised from chick embryos can be drastically reduced if proteinase inhibitors are added to the tissue homogenates. We conclude that the purified 190 kDa dalton protein is a proteolytic fragment of talin. Although markedly reduced by proteinase inhibitors, detection of the 190 kDa protein is not completely abolished, suggesting that some talin may already be cleaved within living cells.
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T
O'Halloran
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K
Burridge
D000818:Animals; D055598:Chemical Phenomena; D002627:Chemistry, Physical; D002642:Chick Embryo; D002645:Chickens; D004591:Electrophoresis, Polyacrylamide Gel; D000939:Epitopes / Q000276:immunology; D005455:Fluorescent Antibody Technique; D005895:Gizzard, Avian; D008970:Molecular Weight; D009124:Muscle Proteins / Q000276:immunology / Q000302:isolation & purification* / Q000378:metabolism; D009130:Muscle, Smooth / Q000032:analysis*; D010446:Peptide Fragments / Q000302:isolation & purification; D016608:Talin; D014018:Tissue Distribution; D016596:Vinculin
D000939:Epitopes; D009124:Muscle Proteins; D010446:Peptide Fragments; D016608:Talin; D016596:Vinculin