Structural characterization of Y1 and Y2 receptors for neuropeptide Y and peptide YY by affinity cross-linking
- Univ. of Michigan, Ann Arbor (USA)
Pharmacological studies indicate that peptide YY (PYY) and neuropeptide Y interact with multiple binding sites, categorized as Y1 and Y2 subtypes. In order to identify and structurally characterize the Y1 and Y2 receptors we covalently cross-linked (125I-Tyr36)PYY to its receptors. The Y2 receptor in rat hippocampus and rabbit kidney membranes was affinity labeled using different homo- and heterobifunctional cross-linking reagents. Analysis by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and autoradiography resulted in a major labeled protein band of Mr = 50,000 in both hippocampal and kidney membranes, which was unaffected by reducing agents. The Y1 receptor was analyzed in membranes from the MC-IXC human neuroblastoma cell line. Autoradiography revealed two labeled bands at Mr = 70,000 and 45,000. As the intensity of the Mr = 45,000 band was reduced by protease inhibitors, it is likely that this band is a degradation product of the larger band. Labeling of these proteins was obtained only when N-5-azido-2-nitrobenzoyloxysuccinimide was employed for cross-linking followed by exposure to UV light. Labeling of the two cross-linked bands was unaffected by reducing agents. The binding of radiolabeled PYY and the intensity of the cross-linked bands, for both the Y1 and Y2 receptors, were inhibited similarly in a dose-dependent manner by increasing concentrations of unlabeled PYY. When exposed to agarose-coupled lectins, the detergent-solubilized Y1 receptor-hormone complex was completely adsorbed by wheat germ agglutinin and partially by ricin communis II. The cross-linked Y2 receptor was almost totally adsorbed by wheat germ agglutinin-agarose and partially adsorbed by concanavalin A. The adsorptions were in all cases blocked by the appropriate hapten sugar.
- OSTI ID:
- 6809865
- Journal Information:
- Journal of Biological Chemistry; (USA), Vol. 265:14; ISSN 0021-9258
- Country of Publication:
- United States
- Language:
- English
Similar Records
Comparison of solubilized and purified plasma membrane and nuclear insulin receptors
Neuropeptide Y receptor binding sites in rat brain: differential autoradiographic localizations with sup 125 I-peptide YY and sup 125 I-neuropeptide Y imply receptor heterogeneity
Related Subjects
63 RADIATION, THERMAL, AND OTHER ENVIRON. POLLUTANT EFFECTS ON LIVING ORGS. AND BIOL. MAT.
PEPTIDES
RECEPTORS
MOLECULAR STRUCTURE
AUTORADIOGRAPHY
BIOCHEMICAL REACTION KINETICS
CELL MEMBRANES
CROSS-LINKING
DOSE-RESPONSE RELATIONSHIPS
ELECTROPHORESIS
ENZYME INHIBITORS
GLYCOPROTEINS
HIPPOCAMPUS
IMIDES
IODINE 125
KIDNEYS
MOLECULAR WEIGHT
PHOTOCHEMISTRY
RABBITS
RATS
REAGENTS
ANIMALS
BETA DECAY RADIOISOTOPES
BODY
BRAIN
CELL CONSTITUENTS
CENTRAL NERVOUS SYSTEM
CHEMICAL REACTIONS
CHEMISTRY
DAYS LIVING RADIOISOTOPES
ELECTRON CAPTURE RADIOISOTOPES
INTERMEDIATE MASS NUCLEI
IODINE ISOTOPES
ISOTOPES
KINETICS
MAMMALS
MEMBRANE PROTEINS
MEMBRANES
NERVOUS SYSTEM
NUCLEI
ODD-EVEN NUCLEI
ORGANIC COMPOUNDS
ORGANIC NITROGEN COMPOUNDS
ORGANS
POLYMERIZATION
PROTEINS
RADIOISOTOPES
REACTION KINETICS
RODENTS
VERTEBRATES
550201* - Biochemistry- Tracer Techniques
560120 - Radiation Effects on Biochemicals
Cells
& Tissue Culture