Identification of the calmodulin-binding components in bovine lens plasma membranes.
Lens membranes, purified from calf lenses, have been labeled by covalent cross-linking to membrane-bound 125I-calmodulin with dithiobis(succinimidyl propionate). Electrophoretic analysis in sodium dodecyl sulfate demonstrated two major 125I-containing products of Mr = 49 000 and 36 000. That the formation of these two components was specifically inhibited by unlabeled calmodulin, or calmodulin antagonists, would indicate that the formation of these components was calmodulin-specific. The size of these two 125I-labeled components was unchanged over a range of 125I-calmodulin or dithiobis(succinimidyl propionate) concentrations indicating that they represent 1:1 complexes between 125I-calmodulin (Mr = 17 000) and Mr-32 000 and Mr-19 000 lens membrane components respectively. Although formation of both cross-linked components exhibited an absolute dependence on Mg2+, the autoradiographic intensity of these components was enhanced when Ca2+ was included with Mg2+ during the cross-linking reaction. Labeling was maximal in 10 mM MgCl2 and approximately 1 microM Ca2+. Treatment of lens membranes with chymotrypsin resulted in the cleavage of MP26 (the major lens membrane protein), with the appearance of a major proteolytic fragment of Mr = 22 000. This proteolysis was not associated with any significant change in either the size or amount of the 125I-calmodulin-labeled membrane components. These results suggest that calmodulin interacts with two membrane proteins, but not significantly with MP26, in the intact lens cell membrane. Our results indicate the need to maintain caution in interpreting direct calcium plus calmodulin effects on MP26 and lens cell junctions.
D000818:Animals; D020346:Aquaporins; D002118:Calcium / Q000494:pharmacology; D002147:Calmodulin / Q000378:metabolism* / Q000494:pharmacology; D002352:Carrier Proteins / Q000032:analysis*; D002417:Cattle; D002462:Cell Membrane / Q000032:analysis; D003459:Crystallins / Q000032:analysis*; D005136:Eye Proteins / Q000378:metabolism; D007365:Intercellular Junctions / Q000187:drug effects; D007457:Iodine Radioisotopes; D007908:Lens, Crystalline / Q000032:analysis*; D008274:Magnesium / Q000494:pharmacology; D008562:Membrane Glycoproteins; D008565:Membrane Proteins / Q000032:analysis*; D008970:Molecular Weight
D020346:Aquaporins; D002147:Calmodulin; D002352:Carrier Proteins; D003459:Crystallins; D005136:Eye Proteins; D007457:Iodine Radioisotopes; D008562:Membrane Glycoproteins; D008565:Membrane Proteins; C030328:aquaporin 0; D008274:Magnesium; D002118:Calcium