Isolation, purification, and characterization of calcitonin gene-related peptide receptor

Intact calcitonin gene-related peptide (CGRP) receptors were solubilized from porcine neural membranes using sodium cholate: potassium buffer. The solubilized receptors were purified sequentially by hydrophobic interaction and ion-exchange chromatography followed by specific affinity chromatography. Using these procedures, we have isolated 2 nmol of highly purified active CGRP receptor to a homogeneity (5 x 10(8)-fold purification). The isolated receptors retained their specificity and the capacity to bind to 125I-CGRP, and showed no cross-reactivity with a number of other peptides, except with amylin having 46% amino acid sequence homology to h-CGRP. The solubilized receptors were adsorbed by WGA-agarose and concanavalin-A, suggesting a glycoprotein nature. SDS-PAGE, size-exclusion HPLC, and autoradiography confirmed that CGRP receptor is a monomeric membrane protein with M(r) 66 kDa.