Chemical crosslinking of a solubilized enkephalin macromolecular complex.

Abstract

Covalently bound [3H]D-Ala2,Met5-enkephalinamide and 125I-labeled D-Ala2,N-Me-Phe4,Met-(O)5-ol-enkephalin macromolecule complexes were prepared by crosslinking the solubilized noncovalent complexes from rat brain. Gel electrophoresis of the partially purified 125I-labeled enkephalin-macromolecule complex under nondenaturing conditions results in a single major radioactive peak. The complex has a Stokes radius of approximately 48 .ANG. as determined by molecular exclusion chromatography; this radius corresponds to a molecular weight of 380,000 for a spherical molecule. In preliminary experiments, sodium dodecyl sulfate electrophoresis of the complex showed a major radioactive peak corresponding to a MW of 35,000. The preparation of these specific covalent enkephalin-macromolecule complexes should be useful in receptor purification and in probing the molecular mechanism of opiate action.