Use of a nitrotryptophan-containing peptide for photoaffinity labeling the pancreatic cholecystokinin receptor

Journal Article · Tue Apr 18 00:00:00 EDT 1989 · Biochemistry; (USA)

DOI:https://doi.org/10.1021/bi00434a047· OSTI ID:5478642

Klueppelberg, U G; Gaisano, H Y; Powers, S P; Miller, L J ^[1]

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1. Mayo Clinic, Rochester, MN (USA)

The authors report the preparation and characterization of a new type of intrinsic photoaffinity labeling probe, on the basis of the incorporation of a photolabile nitrotryptophan into a biologically relevant domain of a peptide. The model system used was the pancreatic cholecystokinin (CCK) receptor, previously affinity labeled with a variety of probes. Those studies have suggested that an M{sub r} = 85,000-95,000 protein is more likely to be labeled as the site of covalent attachment approaches the receptor-binding domain of this hormone. Indeed, CCK has a Trp in the center of its receptor-binding region, and replacement of that residue with 6-nitrotryptophan resulted in a photolabile probe which affinity labeled the same M{sub r} = 85,000-95,000 pancreatic membrane protein. This probe, {sup 125}I-D-Tyr-Gly-((Nle{sup 28,31},6-NO{sub 2}-Trp{sup 30})CCK-26-33), was synthesized by solid-phase and solution techniques and characterized by mass spectrometry. Following oxidative iodination, it was purified on HPLC to 2000 Ci/mmol. Binding to pancreatic membranes was rapid, temperature dependent, reversible, saturable, and specific and was with high affinity. While its binding affinity was only 3-fold lower than that of native CCK-8, this probe was 70-fold less potent than native hormone in stimulating amylase secretion and equally efficacious to native hormone.

• OSTI ID: 5478642
• Journal Information: Biochemistry; (USA), Journal Name: Biochemistry; (USA) Vol. 28:8; ISSN 0006-2960; ISSN BICHA
• Country of Publication: United States
• Language: English

Related Subjects

550201* -- Biochemistry-- Tracer Techniques
59 BASIC BIOLOGICAL SCIENCES
ALKALI METAL COMPOUNDS
AMINO ACIDS
AROMATICS
AZAARENES
AZOLES
BETA DECAY RADIOISOTOPES
BODY
CARBOXYLIC ACIDS
CELL CONSTITUENTS
CELL MEMBRANES
CHEMICAL REACTIONS
CHROMATOGRAPHY
CROSS-LINKING
DAYS LIVING RADIOISOTOPES
DIGESTIVE SYSTEM
ELECTRON CAPTURE RADIOISOTOPES
ENDOCRINE GLANDS
GLANDS
HALIDES
HALOGEN COMPOUNDS
HALOGENATION
HETEROCYCLIC ACIDS
HETEROCYCLIC COMPOUNDS
INDOLES
INORGANIC PHOSPHORS
INTERMEDIATE MASS NUCLEI
IODIDES
IODINATION
IODINE 125
IODINE COMPOUNDS
IODINE ISOTOPES
ISOTOPES
KININS
LABELLING
LIGANDS
LIQUID COLUMN CHROMATOGRAPHY
MEMBRANE PROTEINS
MEMBRANES
MOLECULAR STRUCTURE
NUCLEI
ODD-EVEN NUCLEI
ORGANIC ACIDS
ORGANIC COMPOUNDS
ORGANIC NITROGEN COMPOUNDS
ORGANS
PANCREAS
PEPTIDES
PHOSPHORS
POLYMERIZATION
POLYPEPTIDES
PROTEINS
PYRROLES
RADIOISOTOPES
RECEPTORS
SEPARATION PROCESSES
SODIUM COMPOUNDS
SODIUM IODIDES
TRYPTOPHAN