Neuropeptide Y (NPY) receptors consist of three subtypes, designated NPY, NPY, and NPY. The Y receptor has been cloned. The present study reports the purification of the NPY-Y receptor from porcine brain and its biochemical characterization. NPY receptors were solubilized and purified by sequential hydrophobic interaction, ion exchange, and NPY-affinity chromatography. By use of SDS-polyacrylamide gel electrophoresis, high performance liquid chromatography gel permeation chromatography, and chemical cross-linking studies, the affinity-purified brain NPY-Y receptor was identified as a monomeric glycoprotein with a molecular mass of 60 kDa. Following deglycosylation, the molecular mass of the Y receptor was decreased to 45 kDa. Although the I-NPY binding to the purified NPY receptor was considerably decreased by N-ethylmaleimide, guanine nucleotides had no effect. Therefore, the purified NPY-Y receptor is probably not associated with G-proteins, but may have intramolecular-free sulfhydryl groups. The specific activity of the isolated NPY-Y receptor is 15.8 nmol/mg of protein. The isolated receptor retained its capacity to bind to I-NPY, specific to NPY and peptide YY, and showed no cross-reactivity with any other peptides. Highly purified (10-fold purification) NPY receptor from the brain was identified as the Y subtype as demonstrated by its affinity to C-terminal fragments of NPY, including NPY-(13-36).

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