CHARACTERIZATION OF THE PEPTIDE BINDING REQUIREMENTS FOR THE CLONED HUMAN PANCREATIC POLYPEPTIDE-PREFERRING RECEPTOR

Traditionally, neuropeptide Y (NPY) receptors have been divided into Y 1 and Y2 subtypes based on peptide pharmacology and synaptic localizat ion. Other receptor subtypes have been proposed based on preferences f or NPY, peptide YY (PYY), or pancreatic polypeptide (PP). Recently, we discovered a novel human member of this receptor family exhibiting hi gh affinity for PP and PW. in the current study, we expressed a DNA cl one encoding this human PP-preferring receptor [hPP1 (or Y4)] in Chine se hamster ovary cells and performed a peptide structure-activity stud y. [I-125]pPYY bound to homogenates of hPP1-Chinese hamster ovary cell s with a K-d of 0.064 +/- 0.006 nM and a B-max of 244 +/- 12 fmol/mg p rotein. Human PP inhibited binding with a K-i of 0.023 nM, whereas hum an PW (K-i = 0.31 nM) and human NPY (K-i = 12 nM) were significantly l ess potent. Rat, porcine, and bovine PP inhibited binding with similar affinities to human PP, whereas avian PP was substantially less poten t (K-i = 1 nM). Deletion of the first four amino acids reduced the aff inity of bovine PP to 1 nu. Carboxyl-terminal fragments of NPY and PW also had reduced potency compared with the native peptides. In additio n, deletion of Tyr36-amide produced a substantial reduction in affinit y. Pro34-substituted NPY and PW had modestly increased affinity compar ed with the native peptides, although Gln34-bPP had similar affinity c ompared with bovine PP, The carboxyl-terminally derived Y1 antagonist 1229U91 was a very potent (K-i = 0.042 nM) inhibitor of binding to hPP 1. Thus, the carboxyl-terminal region of PP seems to be the most impor tant part of the peptide for high affinity binding to hPP1. A few key residues (amino acids 2 and 3) in the amino-terminal region of PP cont ribute to the high affinity of the native peptide. Thus, features requ ired for peptide recognition by the hPP1 receptor seem to be distinct from the Y1 and Y2 receptor.