Separate binding sites for intact PTH 1-84 and synthetic PTH 1-34 in canine kidney.
The present studies examine the characteristics of parathyroid hormone (PTH) receptor binding in canine basolateral renal cortical membranes using iodinated preparations of intact bovine PTH 1-84 and [Nle8,Nle18,Tyr34] bPTH 1-34 amide. A solid phase lactoperoxidase technique was used to iodinate the bPTH 1-84. The PTH 1-34 analog was iodinated using chloramine T. Both radioligands were purified by reverse phase high pressure liquid chromatography (HPLC). Specific binding of 125I PTH 1-84 reached equilibrium at 3 hours whereas binding of the 125I PTH 1-84 analog reached equilibrium at 45 minutes. Excess bPTH 1-84 resulted in complete inhibition of binding of 125I bPTH 1-84, whereas 22 +/- 1.6% of the bound radioligand remained bound in the presence of excess synthetic bPTH 1-34. These data suggested the possibility of a binding site for the carboxy-terminal region of intact PTH, or binding sites selective for intact hormone. Therefore, additional studies were performed with PTH fragments, PTH 28-53, PTH 35-84, and PTH 53-84. In contrast to previous studies in other systems, these fragments did not result in significant displacement of 125I PTH 1-84. Analysis of binding of 125I PTH 1-84 and 125I [Nle8,Nle18,Tyr34] PTH 1-34 amide, using LIGAND, both indicated a single site model with similar affinities. Thus, the data are consistent either with multiple receptors with similar affinities or a second binding site for bPTH 1-84 on the same receptor.(ABSTRACT TRUNCATED AT 250 WORDS)
Garcia
J C
JC
Department of Internal Medicine, Washington University School of Medicine, St. Louis, Missouri 63110.
McConkey
C L
CL
Martin
K J
KJ
D000262:Adenylyl Cyclases / Q000378:metabolism; D000818:Animals; D001665:Binding Sites; D001667:Binding, Competitive; D002462:Cell Membrane / Q000648:ultrastructure; D004285:Dogs; D007668:Kidney / Q000166:cytology / Q000201:enzymology / Q000378:metabolism*; D010281:Parathyroid Hormone / Q000378:metabolism*; D010446:Peptide Fragments / Q000378:metabolism*; D011956:Receptors, Cell Surface / Q000648:ultrastructure; D018016:Receptors, Parathyroid Hormone; D019379:Teriparatide; D013997:Time Factors
D010281:Parathyroid Hormone; D010446:Peptide Fragments; D011956:Receptors, Cell Surface; D018016:Receptors, Parathyroid Hormone; D019379:Teriparatide; D000262:Adenylyl Cyclases