Solubilization and Characterization of Calcitonin Gene‐Related Peptide Binding Site from Porcine Spinal Cord

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Abstract
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. l25I-labeled human α-CGRP (125I-CGRP) binding to the solubilized protein was determined by filration using a GF/B glass filter. The maximal binding activity (˜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.