Demonstration of the specific binding of bovine transferrin to the human transferrin receptor in k562 cells: Evidence for interspecies transferrin internalization

Specific binding of ferric bovine transferrin to the human transferrin receptor was investigated using K562 cells propagated in serum‐free medium without transferrin supplemented with 10 −5 elemental iron. Affinity chromatography of solubilized extracts of K562 cells surface‐labeled with 125 I was performed using bovine transferrin‐ and human transferrin‐Sepharose 4B resins. Sodium dodecyl sulfate‐polyacrylamide gel electrophoresis of resin eluates reveal that bovine transferrin specifically binds a M r = 188,000 protein which dissociates into a M r = 94,000 protein under reducing conditions, a finding identical to what is seen with human transferrin. The M r = 94,000 reduced protein isolated by bovine transferrin resin shows an identical one‐dimensional partial proteolytic digestion map with that of the human transferrin receptor. Unlabeled bovine transferrin was shown to specifically compete 125 I‐labeled human transferrin from the human transferrin receptor on the surface of K562 cells at 4°C in a similar manner as unlabeled human transferrin; however, approximately a 2,000‐fold higher concentration of bovine ligand was required to achieve comparable competition (50% inhibition of binding). Indirect immunofluorescence cytolocalization of bovine transferrin in K562 cells grown in serum‐free medium supplemented with ferric bovine transferrin reveal patterns similar to those seen for human transferrin (both focal perinuclear and diffuse cytoplasmic fluorescence). Monensin treatment results in a dramatic accumulation of bovine ligand in perinuclear aggregates, suggesting that it is recycled through the Golgi, as is human transferrin. K562 cells grown in serum‐free medium supplemented with either 300 μg/ml of ferric human or ferric bovine transferrin were found to demonstrate super‐imposable growth curves.