Characterization of a Pachymedusa dacnicolor-Sauvagine analog as a new high-affinity radioligand for corticotropin-releasing factor receptor studies.
The corticotropin-releasing factor (CRF) peptide family comprises the mammalian peptides CRF and the urocortins as well as frog skin sauvagine and fish urophyseal urotensin. Advances in understanding the roles of the CRF ligand family and associated receptors have often relied on radioreceptor assays using labeled CRF ligands. These assays depend on stable, high-affinity CRF analogs that can be labeled, purified, and chemically characterized. Analogs of several of the native peptides have been used in this context, most prominently including sauvagine from the frog Phyllomedusa sauvageii (PS-Svg). Because each of these affords both advantages and disadvantages, new analogs with superior properties would be welcome. We find that a sauvagine-like peptide recently isolated from a different frog species, Pachymedusa dacnicolor (PD-Svg), is a high-affinity agonist whose radioiodinated analog, [(125)ITyr(0)-Glu(1), Nle(17)]-PD-Svg, exhibits improved biochemical properties over those of earlier iodinated agonists. Specifically, the PD-Svg radioligand binds both CRF receptors with comparably high affinity as its PS-Svg counterpart, but detects a greater number of sites on both type 1 and type 2 receptors. PD-Svg is also ∼10 times more potent at stimulating cAMP accumulation in cells expressing the native receptors. Autoradiographic localization using the PD-Svg radioligand shows robust specific binding to rodent brain and peripheral tissues that identifies consensus CRF receptor-expressing sites in a greater number and/or with greater sensitivity than its PS-Svg counterpart. We suggest that labeled analogs of PD-Svg may be useful tools for biochemical, structural, pharmacological, and anatomic studies of CRF receptors.
Clayton Foundation Laboratories for Peptide Biology (M.H.P., J.M.V., K.A.L., C.J.D., C.M., J.E., J.E.R., P.E.S.) and Laboratory of Neuronal Structure and Function (L.A.T., P.E.S.), The Salk Institute for Biological Studies, La Jolla, California.
Marilyn H
MH
Perrin
Clayton Foundation Laboratories for Peptide Biology (M.H.P., J.M.V., K.A.L., C.J.D., C.M., J.E., J.E.R., P.E.S.) and Laboratory of Neuronal Structure and Function (L.A.T., P.E.S.), The Salk Institute for Biological Studies, La Jolla, California.
Laura A
LA
Tan
Clayton Foundation Laboratories for Peptide Biology (M.H.P., J.M.V., K.A.L., C.J.D., C.M., J.E., J.E.R., P.E.S.) and Laboratory of Neuronal Structure and Function (L.A.T., P.E.S.), The Salk Institute for Biological Studies, La Jolla, California.
Joan M
JM
Vaughan
Clayton Foundation Laboratories for Peptide Biology (M.H.P., J.M.V., K.A.L., C.J.D., C.M., J.E., J.E.R., P.E.S.) and Laboratory of Neuronal Structure and Function (L.A.T., P.E.S.), The Salk Institute for Biological Studies, La Jolla, California.
Kathy A
KA
Lewis
Clayton Foundation Laboratories for Peptide Biology (M.H.P., J.M.V., K.A.L., C.J.D., C.M., J.E., J.E.R., P.E.S.) and Laboratory of Neuronal Structure and Function (L.A.T., P.E.S.), The Salk Institute for Biological Studies, La Jolla, California.
Cynthia J
CJ
Donaldson
Clayton Foundation Laboratories for Peptide Biology (M.H.P., J.M.V., K.A.L., C.J.D., C.M., J.E., J.E.R., P.E.S.) and Laboratory of Neuronal Structure and Function (L.A.T., P.E.S.), The Salk Institute for Biological Studies, La Jolla, California.
Charleen
C
Miller
Clayton Foundation Laboratories for Peptide Biology (M.H.P., J.M.V., K.A.L., C.J.D., C.M., J.E., J.E.R., P.E.S.) and Laboratory of Neuronal Structure and Function (L.A.T., P.E.S.), The Salk Institute for Biological Studies, La Jolla, California.
Judit
J
Erchegyi
Clayton Foundation Laboratories for Peptide Biology (M.H.P., J.M.V., K.A.L., C.J.D., C.M., J.E., J.E.R., P.E.S.) and Laboratory of Neuronal Structure and Function (L.A.T., P.E.S.), The Salk Institute for Biological Studies, La Jolla, California.
Jean E
JE
Rivier
Clayton Foundation Laboratories for Peptide Biology (M.H.P., J.M.V., K.A.L., C.J.D., C.M., J.E., J.E.R., P.E.S.) and Laboratory of Neuronal Structure and Function (L.A.T., P.E.S.), The Salk Institute for Biological Studies, La Jolla, California sawchenko@salk.edu.
Paul E
PE
Sawchenko
D000595:Amino Acid Sequence; D029845:Amphibian Proteins / Q000737:chemistry / Q000378:metabolism*; D000818:Animals; D001001:Anura; D002460:Cell Line; D006801:Humans; D007553:Isotope Labeling; D007700:Kinetics; D008024:Ligands; D051379:Mice; D008969:Molecular Sequence Data; D036361:Peptide Hormones / Q000737:chemistry / Q000378:metabolism*; D021381:Protein Transport; D011869:Radioligand Assay / Q000379:methods*; D051381:Rats; D018019:Receptors, Corticotropin-Releasing Hormone / Q000737:chemistry / Q000378:metabolism*
D029845:Amphibian Proteins; D008024:Ligands; D036361:Peptide Hormones; D018019:Receptors, Corticotropin-Releasing Hormone; C025727:sauvagine