Biochemical Characterization and Solubilization of Human NK2Receptor Expressed in Chinese Hamster Ovary Cells

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Abstract
The human ileum neurokinin NK2 receptor has been stably expressed in Chinese hamster ovary (CHO) cells using the dihydrofolate reductase (DHFR) expression system. Amplified cell populations expressing approximately 7×105 NK2 receptors/cell were selected in the presence of the DHFR inhibitor methotrexate. Cross-linking of [125I]NKA to NK2 receptor transfected cells revealed a specifically labeled protein of apparent molecular weight 64 kDa by SDS-polyacrylamide gel electrophoresis. This protein was deglycosylated by the enzymes N-glycosidase F and endoglycosydase F to a protein of apparent molecular weight of 39 kDa. The NK2 receptor was solubilized in an active form from CHO cell membranes using the zwitterionic detergent CHAPS. This method represents a valuable approach for the production of significant amounts of NK2 receptor protein from mammalian cells.