Characterization of the structure of the erythropoietin receptor by ligand blotting

Journal Article · Sat Jun 15 04:00:00 EDT 1991 · Blood (Journal of Hematology); (United States)

OSTI ID:5398612

Atkins, H L; Broudy, V C; Papayannopoulou, T ^[1]

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1. University of Washington, Seattle (USA)

Erythropoietin (Epo) regulates the growth and differentiation of erythroid cells by binding to a specific receptor. We characterized the native Epo receptor on erythroleukemia cell lines by ligand blotting. Solubilized cell membrane proteins were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, transferred onto nitrocellulose, and probed with 125I-Epo. Specificity was demonstrated by inhibition of 125I-Epo binding by unlabeled excess Epo but not other peptide growth factors and by the cellular distribution of the Epo binding protein. A single membrane protein of 61 Kd {plus minus} 4 Kd was sufficient to bind 125I Epo in both human (OCIM2, K562) and murine (GM979, Rauscher, DA-1) cell lines. This finding is consistent with the predicted size of the Epo receptor from the murine cDNA clone. However, chemical crosslinking of 125I-Epo to its receptor has identified two Epo binding proteins of 105 Kd and 85 Kd. This difference may occur because the receptor is size fractionated before Epo binding in the ligand blot, but after Epo binding in crosslinking studies. Ligand blotting demonstrates that the native Epo receptor is composed of a single 61-Kd Epo binding protein, and suggests the presence of additional proteins of 20 to 25 Kd that associate with the receptor after Epo binding.

• OSTI ID: 5398612
• Journal Information: Blood (Journal of Hematology); (United States), Journal Name: Blood (Journal of Hematology); (United States) Vol. 77:12; ISSN BLOOA; ISSN 0006-4971
• Country of Publication: United States
• Language: English

Related Subjects

550201* -- Biochemistry-- Tracer Techniques
59 BASIC BIOLOGICAL SCIENCES
ANIMAL CELLS
ANIMALS
BETA DECAY RADIOISOTOPES
BIOCHEMICAL REACTION KINETICS
BIOLOGICAL MATERIALS
BLOOD
BLOOD CELLS
BODY FLUIDS
CELL CONSTITUENTS
CELL MEMBRANES
CHEMICAL COMPOSITION
CHEMICAL REACTIONS
CROSS-LINKING
DAYS LIVING RADIOISOTOPES
DISEASES
ELECTRON CAPTURE RADIOISOTOPES
ELECTROPHORESIS
ERYTHROCYTES
ERYTHROPOIETIN
GROWTH FACTORS
HORMONES
IMMUNE SYSTEM DISEASES
INTERMEDIATE MASS NUCLEI
INTERNAL CONVERSION RADIOISOTOPES
IODINE 125
IODINE ISOTOPES
ISOTOPE APPLICATIONS
ISOTOPES
KINETICS
LEUKEMIA
LIGANDS
MAMMALS
MATERIALS
MEMBRANE PROTEINS
MEMBRANES
MICE
MITOGENS
MOLECULAR STRUCTURE
MOLECULAR WEIGHT
NEOPLASMS
NUCLEI
ODD-EVEN NUCLEI
ORGANIC COMPOUNDS
PEPTIDE HORMONES
POLYMERIZATION
PROTEINS
RADIOISOTOPES
REACTION KINETICS
RECEPTORS
RODENTS
TRACER TECHNIQUES
TUMOR CELLS
VERTEBRATES