Identification and purification of a kidney membrane protein which specifically binds the amino-terminal domain of native parathyroid hormone.
Nitrocellulose blots of bovine kidney membrane proteins were prepared from denaturing polyacrylamide gels. Strips of the blots were incubated with parathyroid hormone (PTH), washed, and then incubated with antisera against the hormone. Exposure to horseradish peroxidase-linked second antibody led to staining of a 51-kDa protein. No staining was observed in blots not incubated with PTH. Fragments 35-84 and 19-84 of PTH reacted strongly with the antisera, but did not lead to staining of the 51-kDa protein on the blots. Staining was visible, but greatly reduced, when fragment 9-84 was used. Oxidation of the native hormone at positions 8 and 18 led to reductions in staining of the band which were quantitatively similar to the reductions in biological activity induced by such oxidations. These properties suggested that the 51-kDa protein recognizes the amino-terminal portions of PTH, which is the segment of the molecule required for its biological activities. Several micrograms of the 51-kDa protein were purified to homogeneity by selective extraction from the membranes with detergent and by elution from multiple two-dimensional gels. The purified protein retained its PTH-dependent staining and specificity. This protein may be a PTH receptor or a fragment of a PTH receptor from kidney.
D000818:Animals; D002417:Cattle; D002462:Cell Membrane / Q000032:analysis; D004591:Electrophoresis, Polyacrylamide Gel; D007118:Immunoassay; D007124:Immunoenzyme Techniques; D007525:Isoelectric Focusing; D007668:Kidney / Q000032:analysis*; D008565:Membrane Proteins / Q000302:isolation & purification / Q000378:metabolism*; D010084:Oxidation-Reduction; D010281:Parathyroid Hormone / Q000378:metabolism*; D010446:Peptide Fragments / Q000378:metabolism*; D011956:Receptors, Cell Surface / Q000302:isolation & purification / Q000378:metabolism*; D018016:Receptors, Parathyroid Hormone
D008565:Membrane Proteins; D010281:Parathyroid Hormone; D010446:Peptide Fragments; D011956:Receptors, Cell Surface; D018016:Receptors, Parathyroid Hormone