A urokinase receptor-associated protein with specific collagen binding properties

The plasminogen activation cascade system, directed by urokinase and the ur okinase receptor, plays a key role in extracellular proteolysis during tiss ue remodeling, To identify molecular interaction partners of these trigger proteins on the cell, we combined covalent protein cross-linking with mass spectrometry based methods for peptide mapping and primary structure analys is of electrophoretically isolated protein conjugates, A specific tri-molec ular complex was observed upon addition of pro-urokinase to human U937 cell s. This complex included the urokinase receptor, pro-urokinase, and an unkn own, high molecular weight urokinase receptor-associated protein. The trypt ic peptide mixture derived from a cross-linked complex of pro-urokinase and the latter protein was analyzed by nanoelectrospray tandem mass spectromet ric sequencing. This analysis identified the novel protein as the human hom ologue of a murine membrane-bound lectin with hitherto unknown function, Th e human cDNA was cloned and sequenced. The protein, designated uPARAP, is a member of the macrophage mannose receptor protein family and contains a pu tative collagen-binding (fibronectin type II) domain in addition to 8 C-typ e carbohydrate recognition domains. It proved capable of binding strongly t o a single type of collagen, collagen V. This collagen binding reaction at the exact site of plasminogen activation on the cell may lead to adhesive f unctions as well as a contribution to cellular degradation of collagen matr ices.