Vasoactive intestinal peptide receptor on liver plasma membranes: solubilization and cross-linking.
The hepatic receptor for VIP was solubilized from rat liver plasma membranes with 1.4% digitonin and shown to conserve its ability to bind to the ligand. This solubilized receptor demonstrated the high affinity and specificity for VIP (KD approximately 1 nM, binding preference: VIP greater than PHI greater than secretin greater than thymosin alpha 1) which were observed with the nonsolubilized VIP receptor on intact liver plasma membranes. 125I-VIP was next cross-linked to either the solubilized or nonsolubilized receptor using disuccinimido suberate or disuccinimido dithiobis(propionate), and the resulting complexes analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis followed by autoradiography. A broad autoradiographic band which demonstrated a high affinity for VIP was identified at Mr 56,000 (53,000 in the absence of the reducing agent dithiothreitol) for both the solubilized and nonsolubilized receptors. We have thus been able to solubilize from rat liver plasma membranes a receptor with high affinity and specificity for VIP, and confirmed its structural similarity with the native VIP receptor in nonsolubilized membranes using cross-linking techniques.
Nguyen
T D
TD
Department of Medicine, Duke University, Durham, NC.
Kaiser
L M
LM
D000818:Animals; D001345:Autoradiography; D002462:Cell Membrane / Q000378:metabolism; D003432:Cross-Linking Reagents; D004072:Digitonin; D004591:Electrophoresis, Polyacrylamide Gel; D007457:Iodine Radioisotopes; D008099:Liver / Q000378:metabolism*; D008297:Male; D051381:Rats; D011919:Rats, Inbred Strains; D011964:Receptors, Gastrointestinal Hormone / Q000737:chemistry / Q000378:metabolism*; D018005:Receptors, Vasoactive Intestinal Peptide; D012995:Solubility; D014660:Vasoactive Intestinal Peptide / Q000378:metabolism*
D003432:Cross-Linking Reagents; D007457:Iodine Radioisotopes; D011964:Receptors, Gastrointestinal Hormone; D018005:Receptors, Vasoactive Intestinal Peptide; D014660:Vasoactive Intestinal Peptide; D004072:Digitonin