Laminin-nidogen complex. Extraction with chelating agents and structural characterization.
Large quantities of intact laminin-nidogen complex could be extracted from a mouse tumor basement membrane with a physiological buffer containing EDTA. Analysis of the purified complex demonstrated that the two proteins occur in an equimolar ratio and that anchoring of these complexes to the extracellular matrix requires divalent cations. Reversible dissociation of the complex was achieved with 2 M guanidine X HCl and has been used for purification of the individual components. Electron microscopy and binding studies using laminin fragments demonstrated that nidogen interacts specifically with the center of the cross-shaped laminin molecule as represented by the short-arm structure fragment 1. The complex was also useful to confirm and refine a previously proposed dumb-bell structure of nidogen and to prepare and characterize the cell-binding fragment 8 from the long arm of laminin.
D000818:Animals; D001485:Basement Membrane / Q000032:analysis; D001665:Binding Sites; D002118:Calcium / Q000378:metabolism; D002942:Circular Dichroism; D004492:Edetic Acid; D004591:Electrophoresis, Polyacrylamide Gel; D006868:Hydrolysis; D007124:Immunoenzyme Techniques; D007797:Laminin / Q000302:isolation & purification*; D008562:Membrane Glycoproteins; D008565:Membrane Proteins / Q000302:isolation & purification*; D051379:Mice; D008854:Microscopy, Electron; D009374:Neoplasms, Experimental / Q000032:analysis*; D010196:Pancreatic Elastase; D011485:Protein Binding
D007797:Laminin; D008562:Membrane Glycoproteins; D008565:Membrane Proteins; C029967:nidogen; D004492:Edetic Acid; D010196:Pancreatic Elastase; D002118:Calcium