BIOTINYL C-TERMINAL-EXTENDED MOTILIN AS A BIOLOGICALLY-ACTIVE RECEPTOR PROBE

The synthesis, purification, and characterization of biotinylated anal ogues of motilin are reported. The C-terminal of canine motilin was ex tended by the addition of a cysteine residue, and then biotinylated. B iotinyl motilin was purified by following HPLC and characterized by am ino acid analysis. Biotinylation of the ligand was confirmed by ELISA assay with the avidin-biotin system. Biotinyl motilin showed similar a ffinity for binding to rabbit gastric membrane fraction compared to un labeled canine motilin, and also retained functional activity in its a bility to cause contraction of rabbit duodenal segments. To determine the binding of biotinyl motilin in isolated rabbit antral smooth muscl e, cells were incubated with the biotinyl motilin with and without exc ess of unlabeled motilin. Subsequent addition of avidin-biotinylated p eroxidase complex showed the distribution of reaction products over th e cell surface. Bioactive biotinyl motilin provides a useful probe for the demonstration of cell surface motilin receptors and will facilita te receptor purification and characterization.