title: Affinity chromatography demonstrates a direct binding between cytoplasmic dynein and the dynactin complex.
abstract: We used affinity chromatography to probe for a direct binding interaction between cytoplasmic dynein and dynactin. Purified cytoplasmic dynein was found to bind to an affinity column of p150Glued, the largest polypeptide in the dynactin complex. To test the specificity of the interaction, we loaded rat brain cytosol onto the p150Glued affinity column and observed that cytoplasmic dynein from cytosol was specifically retained on the column. Preincubation of the p150Glued affinity matrix with excess exogenous dynein intermediate chain resulted in a significant reduction of dynein binding, suggesting that p150Glued may be interacting with dynein via this polypeptide. Therefore we constructed an affinity column of recombinant dynein intermediate chain and observed that dynactin was retained from rat brain cytosol. These results demonstrate that the native dynein and dynactin complexes are capable of direct in vitro interaction mediated by a direct binding of the dynein intermediate chain to the p150Glued component of the dynactin complex. We have mapped the site of this interaction to the amino-terminal region of p150Glued, which is predicted to form an alpha-helical coiled-coil. Regulation of the dynein-dynactin interaction may prove to be key in the control mechanism for cytoplasmic dynein-mediated vesicular transport.
mesh_terms: D000818:Animals; D001921:Brain / Q000378:metabolism* / Q000648:ultrastructure; D002846:Chromatography, Affinity; D003593:Cytoplasm / Q000378:metabolism; D003600:Cytosol / Q000378:metabolism; D000072159:Dynactin Complex; D004398:Dyneins / Q000737:chemistry / Q000378:metabolism*; D008868:Microtubule Proteins / Q000378:metabolism*; D008869:Microtubule-Associated Proteins / Q000378:metabolism; D011485:Protein Binding; D051381:Rats
chemical_list: D000072159:Dynactin Complex; D008868:Microtubule Proteins; D008869:Microtubule-Associated Proteins; D004398:Dyneins