Direct demonstration of unique mode of natural peptide binding tothe type 2 cholecystokinin receptor using photoaffinity labeling

Maoqing Dong, Laurence J. Miller

Research output: Contribution to journalArticle

3 Scopus citations

Abstract

Direct analysis of mode of peptide docking using intrinsic photoaffinity labeling has provided detailedinsights for the molecular basis of cholecystokinin (CCK) interaction with the type 1 CCK receptor. In thecurrent work, this technique has been applied to the closely related type 2 CCK receptor that also bindsthe natural full agonist peptide, CCK, with high affinity. A series of photolabile CCK analog probes withsites of covalent attachment extending from position 26 through 32 were characterized, with the highestaffinity analogs that possessed full biological activity utilized in photoaffinity labeling. The position 29probe, incorporating a photolabile benzoyl-phenylalanine in that position, was shown to bind with highaffinity and to be a full agonist, with potency not different from that of natural CCK, and to covalently labelthe type 2 CCK receptor in a saturable, specific and efficient manner. Using proteolytic peptide mapping,mutagenesis, and radiochemical Edman degradation sequencing, this probe was shown to establish acovalent bond with type 2 CCK receptor residue Phe120in the first extracellular loop. This was in contrastto its covalent attachment to Glu345in the third extracellular loop of the type 1 CCK receptor, directlydocumenting differences in mode of docking this peptide to these receptors.

Original languageEnglish (US)
Pages (from-to)143-149
Number of pages7
JournalPeptides
Volume46
DOIs
StatePublished - Jan 1 2013

Keywords

  • Cholecystokinin
  • G protein-coupled receptors
  • Ligand binding
  • Photoaffinity labeling
  • Type 2 cholecystokinin receptor

ASJC Scopus subject areas

  • Biochemistry
  • Physiology
  • Endocrinology
  • Cellular and Molecular Neuroscience

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