Purification and functional characterization of the human β 2-adrenergic receptor produced in baculovirus-infected insect cells

• Published in: FEBS letters 1991-05, Vol.282 (2), p.441-444
• Main Authors: Reiländer, Helmut, Boege, Fritz, Vasudevan, Subhash, Maul, Gabi, Hekman, Mirko, Dees, Christian, Hampe, Wolfgang, Helmreich, Ernst J.M., Michel, Hartmut
• Format: Article
• Language: English
• Subjects: Adenylyl Cyclases - metabolism; Affinity chromatography; Alprenolol - metabolism; Baculoviridae; Baculovirus; Cloning, Molecular; Expression; Genetic Vectors; Glycosylation; GTP-Binding Proteins - metabolism; Humans; In Vitro Techniques; Macromolecular Substances; Receptors, Adrenergic, beta - chemistry; Receptors, Adrenergic, beta - isolation & purification; Receptors, Adrenergic, beta - metabolism; Recombinant Proteins - isolation & purification; β-adrenergic receptor
• ISSN: 0014-5793; 1873-3468
• DOI: 10.1016/0014-5793(91)80532-8

A human cDNA fragment bearing the complete coding region for the β 2-adrenergic receptor was introduced into the genome of Autographa california nuclear polyhedrosis virus under the control of the polyhedrin promoter. Binding studies using [ 125I]iodocynnopindolol showed that Sf9 insect cells infected with the recombinant virus expressed ≈ 1 × 10 4 β 2-adrenergic receptors on their cell surface. Photoaffinity labeling of whole cells and membraines revealed a molecular weight of ≈ 46000 for the expressed receptor. The receptor produced in insect cells is glycosylated but the extent and pattern differ from that of the receptor from human tissue. The heterologously expressed receptor was purified by alprenolol affinity chromatography, and was able to activate isolated Gs-protein.