Reduced affinity of lodinated forms of Tyr0 C-type natriuretic peptide for rat natriuretic peptide receptor B

Gaétan Thibault, Kevin L. Grove, Christian F. Deschepper

Research output: Contribution to journalArticlepeer-review

12 Scopus citations


Tyr0CNP is an analogue of C-type natriuretic peptide (CNP) with a tyrosine residue added to the NH2 terminus to allow its iodination. In the present study, the suitability of iodinated Tyr0CNP as a ligand was tested, and its potency was compared with that of other natural rat natriuretic peptides or structural analogues by radioligand binding experiments. Binding studies were performed on membranes of COS-1 cells transfected with expression plasmids for either rat natriuretic peptide receptor (NPR)-A, rat NPR-B, or bovine NPR-C. 125I-ANP(99-126) was used as a ligand to assess the binding characteristics of NPR-A and -C, and 125I-Tyr0CNP was used to study NPR-B. Binding associated to membranes of nontransfected COS cells was always <3% of the total binding observed in membranes from cells transfected with receptor expression plasmids. Receptor densities in transfected cells ranged from 500 to 2500 fmol/mg of protein. High performance liquid chromatography and ionspray mass spectrometry analyses revealed that the reagents used in the course of iodination (lactoperoxidase, chloramine T, or N-chloromorpholine) altered the structure of Tyr0CNP, most likely by changing thethiolof the Met17 residue into a sulfoxide. To further evaluate the usefulness of forms of iodinated Tyr0CNP as radioligands, we performed iodination of the peptide with cold iodine (Na-127I-). After purification by high performance liquid chromatography, three different modified peptides (i.e., Tyr0Met(O)17CNP, 127I-Tyr°Met(O)17CNP, and 127I-Tyr0Met(O)17CNP) were recovered, and they were compared with CNP-22, Tyr0CNP, ANP(99-126), BNP-32, and des[Gln18,Ser19,Gly20,Leu21,Gly 22]ANP(4-23) NH2 (c-ANP) for their ability to bind to transfected receptors. The binding affinity of Tyr0CNP for NPR-A and -B receptors is similar to that of CNP. However, oxidation of the Met17 residue into methionine sulfoxide reduces the affinity of the compound for NPR-B by > 10-fold, whereas the addition of one or two iodines did not further reduce its affinity. Similar results were obtained on evaluation of the ability of the oxidized form of monoiodinated Tyr0CNP on the cGMP responses in cells transfected with NPR-B. In conclusion, the suitability of iodinated forms of Tyr0CNP as radioligands for binding studies on rat NPR-B is not optimal, and the results of studies using such compounds for the detection, identification, and quantification of this receptor should be interpreted with caution.

Original languageEnglish (US)
Pages (from-to)1046-1053
Number of pages8
JournalMolecular pharmacology
Issue number6
StatePublished - Dec 1995
Externally publishedYes

ASJC Scopus subject areas

  • Molecular Medicine
  • Pharmacology


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