Solubilization and characterization of calcitonin gene-related peptide binding site from porcine spinal cord

• Published in: Journal of neurochemistry Vol. 50; no. 2; p. 480
• Main Authors: Hiroshima, O, Sano, Y, Yuzuriha, T, Yamato, C, Saito, A, Okamura, N, Uchiyama, Y, Kimura, S, Goto, K
• Format: Journal Article
• Language: English
• Published: England 01.02.1988
• Subjects: Animals; Binding Sites; Calcitonin Gene-Related Peptide; Cell Membrane - metabolism; Cholic Acids; Chromatography, Gel; Hydrogen-Ion Concentration; Molecular Weight; Neuropeptides - metabolism; Receptors, Calcitonin; Receptors, Cell Surface - isolation & purification; Receptors, Cell Surface - metabolism; Solubility; Spinal Cord - metabolism; Swine; Temperature
• ISSN: 0022-3042
• DOI: 10.1111/j.1471-4159.1988.tb02936.x

The binding site for calcitonin gene-related peptide (CGRP) was solubilized with 3-[(3-cholamidopropyl)dimethylammonio]-1-propane sulfonate (CHAPS) in an active form from porcine spinal cord. 125I-labeled human alpha-CGRP (125I-CGRP) binding to the solubilized protein was determined by filtration using a GF/B glass filter. The maximal binding activity (approximately 60% of the crude membrane fraction) was obtained with 5 mM CHAPS. 125I-CGRP binding to the solubilized protein was of high affinity, saturability, and high specificity, having KD and Bmax values of 3.69 pM and 338 fmol/mg of protein, respectively. The binding activity was eluted in a single peak with a molecular mass of 400,000 daltons by gel filtration on TSK gel G4000SW. These results suggest that the solubilized protein may be responsible for the specific binding site.