Identification of 125I-labeled rat reticulocyte membrane proteins with affinity for transferrin.

The present studies were performed to identify reticulocyte membrane receptors for transferrin. Rat reticulocytes were labeled in vitro with 1251 via lactoperoxdose; cell ghosts were then prepared and solubilized with Triton X-100. Binding of soluble membrane1251 components to transferrin conjugated to Sepharose (Seph. tfn) was three times greater than to albumin conjugated to Sepharose (Sephalb). Soluble membrane-’25l components released from Seph-tfn by 8 M urea rebound to Seph-tfn but not to Seph-alb. Polyacrylamide gel electrophoresis (PAGE) of membrane components released from Seph-tfn showed three proteins with estimated molecular weights of 76,000, 95,000, and 145,000 daltons. Antibody to transferrin formed a precipitate upon incubation with soluble reticulocyte membrane components but not with soluble membrane components from erythrocytes or reticulocytes exposed to trypsin. PAGE of precipitates formed by incubation of antitransferrin and soluble 125 l-reticulocyte membrane components showed three nonantibody proteins and corresponding 1251 peaks with molecular weights of 76,000, 95,000, and 145,000 daltons. All three proteins increased in quantity when the amount of transferrin was increased in the immune precipitate by addit on of “cold” exogenous transferrin, but only the 95,000and 45,000-dalton components had increased ‘25I. These data indicate that two reticulocyte membrane components (95,000 and 145,000 daltons) can be iodinated and shown to have affinity for transferrin (76,000 daltons). The lack of components with such properties in membranes from erythrocytes or trypsinized reticulocytes, cells lacking the ability to bind transferrin, suggests that the two reticulocyte membrane proteins function as the transferrin receptor.