[36] Purification of the LDL receptor

This chapter describes the methods for the purification of the low-density lipoprotein (LDL) receptor from various sources. These methods are designed to enable researchers to obtain chemical amounts of purified receptor from a rich source—the adrenal cortex—and smaller amounts of radiolabeled LDL receptors from less abundant sources, including cultured cells. The LDL receptor on the surface of mammalian cells binds plasma LDL—a cholesterol transport protein—and, thereby initiates a chain of events culminating in the internalization of the LDL with concomitant delivery of cholesterol to the cell. In mediating the endocytosis of LDL, the LDL receptor migrates continuously from one cell organelle to another. After synthesis in the rough endoplasmic reticulum, the receptor travels to the Golgi complex and then to random sites on the plasma membrane. Within minutes, it binds LDL and clusters with other receptors in clathrin-coated pits. The receptor–LDL complex is internalized in coated vesicles, which rapidly shed their clathrin coat and fuse with other vesicles to form endosomes. Within the acidic environment of the endosome, the receptor and LDL separate—the receptor returns to the surface and the LDL proceeds to the lysosome, where it is degraded, yielding its cholesterol for cellular metabolism. After returning to the surface, the receptor binds another LDL particle and, thus initiates another cycle of endocytosis.