title: Developmental changes in insulin receptors of pig red blood cells.
abstract: Scatchard analysis of the insulin binding to pig reticulocytes, fetal red cells, and adult erythrocytes showed the maximum number of high-affinity binding sites per cell to be 274, 147, and 29, respectively. All three cell types displayed a practically identical dissociation constant of approximately 1.22 X 10(-8) M at the high-affinity region. A long-term in vitro incubation of the fetal red cells and reticulocytes under tissue culture conditions was accompanied by a significant loss of insulin-binding capacity without any appreciable alteration of the dissociation constant. The isolation and characterization of insulin-receptor complexes from these cell types were carried out to establish whether the difference in insulin-binding capacity was due to the difference in the amount of the same species or due to different species of insulin receptors. Membrane proteins were extracted with Triton X-102 and fractionated by DEAE-Sephacel ion-exchange column chromatography. Each peak sample was complexed with 125I-insulin, and the complexes were covalently crosslinked and then applied to a Sepharose CL-6B column. A 95,000-Da complex was obtained from adult pig erythrocyte membranes; 220,000- and and 95,000-Da complex was obtained from adult pig erythrocyte membranes; 220,000- and 95,000-Da complexes from pig reticulocyte membranes; and greater than 600,000-, 220,000-, and 95,000-Da complexes from pig fetal cell membranes. Upon sodium dodecyl sulfate-polyacrylamide gel electrophoresis under a nonreducing condition, the 95,000-Da complex was dissociated into a 53,000-Da component; the greater than 600,000-Da complex into greater than 320,000-, 130,000-, and 53,000-Da components; and the 220,000-Da complex was dissociated into 220,000-, 130,000-, and 53,000-Da components. These findings strongly suggest that the decrease in insulin binding during the developmental changes of red blood cells is due to a disappearance of high-molecular-weight insulin receptors rather than a decrease in the amount of the smaller receptor molecules.
mesh_terms: D000375:Aging; D000818:Animals; D000831:Animals, Newborn; D004905:Erythrocyte Aging; D004910:Erythrocyte Membrane / Q000378:metabolism; D004912:Erythrocytes / Q000378:metabolism*; D005312:Fetal Blood / Q000378:metabolism; D066298:In Vitro Techniques; D011485:Protein Binding; D011972:Receptor, Insulin / Q000378:metabolism*; D012156:Reticulocytes / Q000378:metabolism; D012995:Solubility; D013552:Swine
chemical_list: D011972:Receptor, Insulin