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.
Wimalawansa
S J
SJ
Department of Chemical Pathology, Royal Postgraduate Medical School, London, UK.
Gunasekera
R D
RD
Zhang
F
F
D000818:Animals; D015740:Calcitonin Gene-Related Peptide / Q000378:metabolism; D002531:Cerebellum / Q000737:chemistry*; D002853:Chromatography, Liquid; D003902:Detergents; D004591:Electrophoresis, Polyacrylamide Gel; D007700:Kinetics; D018015:Receptors, Calcitonin Gene-Related Peptide / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D012995:Solubility; D013552:Swine; D014867:Water / Q000737:chemistry
D003902:Detergents; D018015:Receptors, Calcitonin Gene-Related Peptide; D014867:Water; D015740:Calcitonin Gene-Related Peptide