Identification and characterization of cross-link products of E- and N-cadherin contained in PLC cells and enriched after IP. (A and B) Total cytoskeletal proteins of PLC cells taken up in RIPA buffer were reacted with cross-link (CL) reagents (in this case oxidative disulfide bridge formation in the presence of Cu²⁺-phenanthroline) and subjected to IP with antibodies to E-cadherin (E-cad; A) or N-cadherin (N-cad; B). The pelleted immunoprecipitates were then analyzed by SDS-PAGE under nonreducing conditions, and polypeptides were identified by immunoblotting using antibodies to E (lane E)- or N (lane N)-cadherin. Spn, remaining supernatant; pellet, material contained in the final sediment. Note that in both kinds of immunoprecipitates the occurrence of a large proportion of E- and N-cadherin in a common polypeptide with an apparent molecular mass of ∼230 kD (molecular mass markers analyzed in parallel are indicated on the left margin). Note also the formation of some proteolytic breakdown products in the specific immunoprecipitates. (C) Cross-link products of higher molecular masses up to ∼450 and >600 kD can be obtained with the very long distance (>11 Å) lysine–lysine cross-link reagent BS³. Positions of major monomeric and cross-linked molecules are marked by dots. White lines indicate that intervening lanes have been spliced out.