Method for isolation of kappa-opioid binding sites by dynorphin affinity chromatography
- 1 April 1990
- journal article
- research article
- Published by Wiley in Journal of Neuroscience Research
- Vol. 25 (4) , 549-555
- https://doi.org/10.1002/jnr.490250412
Abstract
A kappa‐opioid receptor subtype was purified from a digitonin extract of frog brain membranes, using affinity chromatography. The affinity resin was prepared by coupling dynorphin (1–10) to AH Sepharose 4B. The purified receptor binds 4,750 pmol [3H]ethylketocyclazocine (EKC) per mg protein (5,600‐fold purification over the membrane‐bound receptor) with a Kd of 9.1 nM. The addition of cholesterol‐phosphatidylethanolamine (2:1) enhanced 3.6‐fold the binding activity of the purified material, which gives a purification very close to the theoretical. The purified receptor protein exhibits high affinity for kappa‐selective ligands. The purified fraction shows one major band (65,000 Mr) in sodium dodecyl sulfate (SDS) gel electrophoresis.Keywords
This publication has 27 references indexed in Scilit:
- Lipid Requirement for μ Opioid Receptor BindingJournal of Neurochemistry, 1987
- Purification to apparent homogeneity of a mu-type opioid receptor from rat brain.Proceedings of the National Academy of Sciences, 1986
- k‐Binding and Degradation of [3H]Dynorphin A (1–8) and [3H]Dynorphin A (1–9) in Suspensions of Guinea Pig Brain MembranesJournal of Neurochemistry, 1985
- Solubilization and characterization of mu, delta, and kappa opioid binding sites from guinea pig brain: physical separation of kappa receptors.Proceedings of the National Academy of Sciences, 1984
- Isolation of opiate binding components by affinity chromatography and reconstitution of binding activities.Proceedings of the National Academy of Sciences, 1983
- A rapid filtration assay for soluble receptors using polyethylenimine-treated filtersAnalytical Biochemistry, 1983
- Dynorphin1–8 and dynorphin1–9 are ligands for the κ-subtype of opiate receptorNature, 1982
- CHARACTERIZATION OF THE k‐SUBTYPE OF THE OPIATE RECEPTOR IN THE GUINEA‐PIG BRAINBritish Journal of Pharmacology, 1981
- The Synthesis of [4‐Carboranylalanine, 5‐Leucine]‐Enkephalin (Including an Improved Preparation of t‐Butoxycarbonyl‐L‐o‐carboranylalnine, New Derivatives of L‐Propargylglycine, and a Note on Melanotropic and Opiate Receptor Binding Characteristics)Helvetica Chimica Acta, 1979
- A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye bindingAnalytical Biochemistry, 1976