AJP - Gastrointestinal and Liver Physiology, Vol 256, Issue 3 637-G643, Copyright © 1989 by American Physiological Society
Cross-linking of neuropeptide Y to its receptor on rat brain membranes
P. J. Mannon, I. L. Taylor, L. M. Kaiser and T. D. Nguyen
Department of Medicine, Duke University Medical Center, Durham, North Carolina 27710.
The receptor for neuropeptide Y (NPY) was identified on rat brain membranes
after covalent labeling with 125I-NPY using the homobifunctional
cross-linkers disuccinimido suberate and disuccinimido
dithiobis(propionate) and the heterobifunctional photoactive cross-linker
succinimido 4-azidobenzoate. Analysis by sodium dodecyl
sulfate-polyacrylamide gel electrophoresis followed by autoradiography
revealed the presence of two bands at Mr 62,000 and 39,000. Both species
showed the same high affinity for 125I-NPY. Exposure to reducing agents did
not change the migration of these bands. When the NPY receptor complex was
solubilized from the membranes with 1% Triton X-100 and analyzed by gel
filtration chromatography, it eluted from a Fractogel TSK 55F column as a
peak at approximately 65 kDa. This peak was asymmetric with a shoulder of
radioactivity that probably reflects the smaller receptor species. These
data indicate that the NPY receptor on rat brain membranes is a monomeric
58-kDa unit (62 kDa minus the mass of the cross-linked NPY) without
covalently or noncovalently linked subunits. The smaller 39-kDa species may
be an immature form of the 62-kDa species, a second distinct receptor, or a
degradation product of the 62-kDa band.
