Characterization of receptors for VIP on pancreatic acinar cell plasma membranes using covalent cross-linking.
Vasoactive intestinal peptide (VIP) receptors on guinea pig pancreatic acini differ from those on all other tissues in containing a high-affinity VIP receptor and a low-affinity VIP receptor that has a high affinity for secretin. To characterize the molecular components of these receptors, 125I-VIP was covalently cross-linked to these receptors by four different cross-linking agents: disuccinimidyl suberate, ethylene glycol bis (succinimidyl succinate), dithiobis (succinimidylpropionate), and m-maleimidobenzoyl N-hydroxysuccinimide ester. Analysis by sodium dodecyl sulfate-polyacrylamide gel electrophoresis demonstrated a single major polypeptide band of Mr 45,000 and a minor polypeptide band of Mr 30,000 were cross-linked to 125I-VIP. Covalent cross-linking only occurred when a cross-linking agent was added, was inhibited by GTP, was inhibited by VIP receptor agonists or antagonists that interact with VIP receptors, and not by other pancreatic secretagogues that interact with different receptors. For inhibiting both cross-linking and binding of 125I-VIP to the major polypeptide Mr 45,000 and the minor polypeptide Mr 30,000 components, the relative potencies were VIP greater than helodermin greater than rat growth hormone releasing factor greater than peptide histidine isoleucine greater than secretin. The apparent molecular weight of the cross-linked polypeptides were unchanged by dithiothreitol. Thus the high-affinity VIP receptor on pancreatic acinar cell membranes consists of a single major polypeptide of Mr 45,000, and this polypeptide is not a subunit of a larger disulfide-linked structure. Furthermore, either the low-affinity VIP/secretin-preferring receptor was not covalently cross-linked under the experimental conditions or it consists of a major polypeptide with the same molecular weight as the high-affinity VIP receptor.
D000818:Animals; D001839:Bombesin / Q000494:pharmacology; D002217:Carbachol / Q000494:pharmacology; D002462:Cell Membrane / Q000378:metabolism; D003432:Cross-Linking Reagents; D013007:Growth Hormone-Releasing Hormone / Q000494:pharmacology; D006160:Guanosine Triphosphate / Q000494:pharmacology; D006168:Guinea Pigs; D066298:In Vitro Techniques; D036341:Intercellular Signaling Peptides and Proteins; D008970:Molecular Weight; D010179:Pancreas / Q000378:metabolism*; D010451:Peptide PHI / Q000494:pharmacology; D010455:Peptides / Q000494:pharmacology; D011964:Receptors, Gastrointestinal Hormone / Q000378:metabolism*; D018005:Receptors, Vasoactive Intestinal Peptide; D012633:Secretin / Q000378:metabolism; D012844:Sincalide / Q000494:pharmacology; D013373:Substance P / Q000494:pharmacology; D014660:Vasoactive Intestinal Peptide / Q000378:metabolism*
D003432:Cross-Linking Reagents; D036341:Intercellular Signaling Peptides and Proteins; D010451:Peptide PHI; D010455:Peptides; D011964:Receptors, Gastrointestinal Hormone; D018005:Receptors, Vasoactive Intestinal Peptide; D012633:Secretin; D013373:Substance P; D014660:Vasoactive Intestinal Peptide; D006160:Guanosine Triphosphate; C040442:heliodermin; D002217:Carbachol; D013007:Growth Hormone-Releasing Hormone; D012844:Sincalide; D001839:Bombesin