Biotinylated human. beta. -endorphins as probes for the opioid receptor

Journal Article · Tue Jan 05 04:00:00 EST 1988 · J. Biol. Chem.; (United States)

OSTI ID:6922187

Hochhaus, G; Gibson, B W; Sadee, W

The reaction of human ..beta..-endorphin and biotinyl N-hydroxysuccinimide with or without spacer arm, afforded a series of products that were separated by high performance liquid chromatography (HPLC). Liquid secondary ion mass spectrometry of the biotinylated products and their tryptic digests produced abundant protonated molecular ions (MH/sup +/), which specified the number and location of biotinylation. Between 1 and 4 biotinyl residues were incorporated per human ..beta..-endorphin molecule, at Lys-9, -19, -24, -28, and -29, but not at the amino-terminal Try-1. Three HPLC fractions were isolated for receptor binding studies monobiotinylation of Lys-9, Lys-19, and a mixture of Lys-24, Lys-28, and Lys-29 derivatives. IC/sub 50/ values for binding to ..mu.. and delta opioid receptor sites were 3-8 times higher for monobiotinylated derivatives than for the parent human ..beta..-endorphin. Association with avidin decreased opioid receptor affinities for the C/sub 6/ spacer derivative biotinylated at position Lys-9, which is close to the (1-5) enkephalin receptor region. In contrast, avidin did not affect or even increased apparent affinities to ..mu.. and delta sites for derivatives biotinylated at the ..cap alpha..-helical part of the molecule (Lys-19, -24, -28, and -29). Biotinylated human ..beta..-endorphins also bound to low affinity nonopioid binding sites on NG-108-15 cells; however, affinities to these sites were considerably reduced when derivatives were bound to avidin. The ability of biotinylated human ..beta..-endorphin to cross-link the ..mu.. and delta opioid receptors to avidin allows application of the biotin-avidin system as a molecular probe of the opioid receptor.

• Research Organization: Univ. of California, San Francisco (USA)
• OSTI ID: 6922187
• Journal Information: J. Biol. Chem.; (United States), Journal Name: J. Biol. Chem.; (United States) Vol. 263:1; ISSN JBCHA
• Country of Publication: United States
• Language: English

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63 RADIATION, THERMAL, AND OTHER ENVIRON. POLLUTANT EFFECTS ON LIVING ORGS. AND BIOL. MAT.
ANESTHETICS
ANIMALS
ANTICONVULSANTS
AROMATICS
AZINES
BARBITURATES
BIOLOGICAL EFFECTS
BODY
CARCINOGENESIS
CELL CONSTITUENTS
CENTRAL NERVOUS SYSTEM DEPRESSANTS
CONDENSED AROMATICS
DIGESTIVE SYSTEM
DRUGS
ENZYMES
GLANDS
HETEROCYCLIC COMPOUNDS
HYDROCARBONS
HYDROLASES
HYPNOTICS AND SEDATIVES
LIVER
MAMMALS
METABOLISM
MICROSOMES
ORGANIC COMPOUNDS
ORGANIC NITROGEN COMPOUNDS
ORGANIC OXYGEN COMPOUNDS
ORGANOIDS
ORGANS
PATHOGENESIS
PHENOBARBITAL
POLYCYCLIC AROMATIC HYDROCARBONS
PYRIMIDINES
RACEMIZATION
RATS
RODENTS
TRIPHENYLENE
VERTEBRATES