Binding and endocytosis of 39 kDa protein by mdbk cells

A 39 kDa protein copurifies with the low density lipoprotein receptor‐related protein (LRP) and regulates ligand interactions with LRP. In our recent studies on the clearance of the 39 kDa protein in vivo, we demonstrated that once the liver LRP receptors were saturated, the kidney necame the major organ responsible for the 39 kDa protein clearance (Warshawsky et al., 1993, J. Clin. Invest., 92:937‐944). The current study was undertaken in order to investigate the potential binding and cellular processing of the 39 kDa protein by kidney‐derived MDBK cells. Herein we demonstrate specific, high‐affinity, saturable, and Ca²⁺‐dependent binding of the ¹²⁵I‐39 kDa protein to MDBK cells (K_d ∼ 10‐15 nM, 50‐70,000 bindings sites per cell). Cellular uptake and degradation of the ¹²⁵I‐39 kDa protein by MDBK cells was also demonstrated with kinetics typical of receptor‐mediated endocytosis. Using chemical crosslinking we show that LRP in part mediates the binding of ¹²⁵I‐39 kDa protein to the MDBK cell surface. In addition, the presence of functional LRP on the MDBK cell surface was confirmed by the specific binding of activated α₂‐macroglobulin, another ligand of LRP. Our data thus demonstrate the ability of kidney‐derived MDBK cells to specifically bind, endocytose, and degrade the 39 kDa protein. © 1995 Wiley‐Liss, Inc.