Molecular architecture of secretin receptors: the specific covalent labelling of a 51 kDa peptide after cross-linking of [125I]iodosecretin to intact rat pancreatic acini.
p-Azidophenylglyoxal (APG), a heterobifunctional reagent with one group reacting selectively with arginine residues and another group photoactivable, was used to cross-link [125I]secretin prebound to intact rat pancreatic acini. The best yield was obtained when the [125I]secretin-acini complex was incubated under dim light with 2 mM APG at 37 degrees C and pH 8.0, followed by photolysis at 312 nm. The main secretin binding peptide cross-linked under reducing conditions, when tested by SDS-PAGE and autoradiography: (i) had a molecular mass of 51 kDa and was not a subunit of a larger disulfide-linked structure, and (ii) was distinct from the main VIP binding peptide coexisting in the same preparation.
Gossen
D
D
Department of Biochemistry and Nutrition, Medical School, Université Libre de Bruxelles, Belgium.
Poloczek
P
P
Svoboda
M
M
Christophe
J
J
D000818:Animals; D001386:Azides / Q000494:pharmacology; D001667:Binding, Competitive; D003432:Cross-Linking Reagents / Q000494:pharmacology; D066298:In Vitro Techniques; D007457:Iodine Radioisotopes; D008297:Male; D008970:Molecular Weight; D010179:Pancreas / Q000378:metabolism*; D010658:Phenylglyoxal / Q000031:analogs & derivatives / Q000494:pharmacology; D010777:Photochemistry; D051381:Rats; D011919:Rats, Inbred Strains; D043562:Receptors, G-Protein-Coupled; D011964:Receptors, Gastrointestinal Hormone / Q000378:metabolism*; D012633:Secretin / Q000378:metabolism*
D001386:Azides; D003432:Cross-Linking Reagents; D007457:Iodine Radioisotopes; D043562:Receptors, G-Protein-Coupled; D011964:Receptors, Gastrointestinal Hormone; C029937:secretin receptor; D012633:Secretin; C032404:4-azidophenylglyoxal; D010658:Phenylglyoxal