Characterization of the binding of bovine thrombin to isolated rat hepatocytes.
Isolated rat hepatocytes possess per cell 4,500 high-affinity binding sites for thrombin with a Kd of 30-40 pM, and 2.8 X 10(5) low-affinity sites with a Kd of 30 nM. These binding sites are highly specific for thrombin. Half-maximal binding of 125I-labelled thrombin is achieved after 3 min at 37 degrees C and 7 min at 4 degrees C. The reversibly bound fraction of the ligand dissociates according to a biexponential time course with the rate constants 1-2 X 10(-2) s-1 and 3-4 X 10(-4) s-1. Part of the tracer remains cell-associated even after prolonged incubation, but all cell-associated radioactivity migrates as intact thrombin upon sodium dodecyl sulphate polyacrylamide gel electrophoresis. The bound thrombin is minimally endocytosed as judged by the resistance to pH 3-treatment. Cell-associated radioactivity dissociated from the cells binds just as well in a receptor assay as tracer incubated in a conditioned medium under the same conditions, indicating the absence of a quantitatively important receptor-mediated degradation of the ligand.
Weyer
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B
Institute of Physiology, University of Aarhus, Denmark.
Petersen
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TE
Sonne
O
O
D000818:Animals; D002417:Cattle; D002478:Cells, Cultured; D004705:Endocytosis; D006863:Hydrogen-Ion Concentration; D008099:Liver / Q000166:cytology / Q000378:metabolism*; D008297:Male; D051381:Rats; D011956:Receptors, Cell Surface / Q000378:metabolism; D018179:Receptors, Thrombin; D013917:Thrombin / Q000378:metabolism*
D011956:Receptors, Cell Surface; D018179:Receptors, Thrombin; D013917:Thrombin