Separation and identification of the major constituents of cytoplasmic gels from macrophages.
Proteins which bind to actin filaments in macrophages were investigated by developing a procedure for the isolation of cytoplasmic gels. The gels were found to consist of five major constituents: actin, filamin and the 105-kDa, 70-kDa and 55-kDa components. Prolonged exposure of this macromolecular complex to high-ionic-strength buffer solubilized almost all the proteins, leaving behind the 55-kDa component along with a large amount of actin. Gel filtration of the solubilized extract led to the isolation of five constituents comprising actin, filamin, the 105-kDa and 70-kDa polypeptides, plus a molecular species which eluted at the position of a 280-kDa globular protein. The biochemical and immunological properties of the 105-kDa component were analogous to those of alpha-actinin. Although several attempts were made to correlate the three other constituents (280-kDa, 70-kDa and 55-kDa) with known cytoskeletal proteins, their identity remains to be established. alpha-Actinin, and the 280-kDa and 70-kDa species all exhibited the ability to co-sediment with F-actin and to pack actin filaments into bundles. The bundling activity of the 70-kDa protein was significantly decreased in the presence of micromolar concentrations of calcium, while the activity of the 280-kDa protein was not. Such a Ca2+-sensitive protein could be very important in controlling the local cytoplasmic viscosity.
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D000199:Actins / Q000302:isolation & purification; D000818:Animals; D002118:Calcium; D002352:Carrier Proteins / Q000302:isolation & purification*; D003593:Cytoplasm / Q000032:analysis; D005782:Gels; D007120:Immunochemistry; D008264:Macrophages / Q000032:analysis*; D008274:Magnesium; D008562:Membrane Glycoproteins; D008565:Membrane Proteins; D008840:Microfilament Proteins; D008854:Microscopy, Electron; D008970:Molecular Weight; D011650:Pulmonary Alveoli / Q000032:analysis; D011817:Rabbits; D014746:Vimentin
D000199:Actins; D002352:Carrier Proteins; D005782:Gels; D008562:Membrane Glycoproteins; D008565:Membrane Proteins; D008840:Microfilament Proteins; D014746:Vimentin; C026742:plastin; D008274:Magnesium; D002118:Calcium