Characterization of the human Y1 neuropeptide Y receptor expressed in insect cells

Molecular and Cellular Endocrinology
M MunozP Walker

Abstract

We have expressed the human Y1 NPY receptor in insect cells using a recombinant baculovirus (BacY1). Non-linear curve fitting of competition binding data indicates the presence of 500,000-750,000 saturable NPY binding sites per cell. The affinity of the recombinant Y1 receptor for NPY (Kd = 0.38 +/- 0.8 nM) was identical to the natural receptor. We used a foreign epitope to characterize, immunopurify, and localize the recombinant protein. Cross-linking experiments identified a 65 kDa band as the major NPY binding species. Confocal microscopy indicated that although some recombinant proteins are detectable as early as 12 h post-infection, significant expression at the cell surface is only seen 24-48 h post-infection. We also describe a procedure to treat infected Sf21 cells in such a way that they can be frozen and stored at -80 degrees C for many months before being used for binding studies.

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Citations

Mar 26, 2002·Molecular and Cellular Endocrinology·Ulrica MarklundA Jonas Ekstrand
Dec 10, 1997·Journal of Chromatography. B, Biomedical Sciences and Applications·K Löster, D Josić
Jan 8, 2005·Phytomedicine : International Journal of Phytotherapy and Phytopharmacology·E A AbourashedA Brattström
Jul 13, 2011·Journal of Receptor and Signal Transduction Research·Nathalie PopArmin Buschauer
Mar 11, 2003·Experimental Biology and Medicine·Magnus M BerglundDonald R Gehlert

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