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GPCR Engineering Yields High-Resolution Structural Insights into β2-Adrenergic Receptor Function
Daniel M. Rosenbaum,1*Vadim Cherezov,2*Michael A. Hanson,2Søren G. F. Rasmussen,1Foon Sun Thian,1Tong Sun Kobilka,1Hee-Jung Choi,1,3Xiao-Jie Yao,1William I. Weis,1,3Raymond C. Stevens,2Brian K. Kobilka1
The β2-adrenergic receptor (β2AR) is a well-studiedprototype for heterotrimeric guanine nucleotide–bindingprotein (G protein)–coupled receptors (GPCRs) that respondto diffusible hormones and neurotransmitters. To overcome thestructural flexibility of the β2AR and to facilitate itscrystallization, we engineered a β2AR fusion protein inwhich T4 lysozyme (T4L) replaces most of the third intracellularloop of the GPCR ("β2AR-T4L") and showed that this proteinretains near-native pharmacologic properties. Analysis of adrenergicreceptor ligand-binding mutants within the context of the reportedhigh-resolution structure of β2AR-T4L provides insightsinto inverse-agonist binding and the structural changes requiredto accommodate catecholamine agonists. Amino acids known toregulate receptor function are linked through packing interactionsand a network of hydrogen bonds, suggesting a conformationalpathway from the ligand-binding pocket to regions that interactwith G proteins.
1 Department of Molecular and Cellular Physiology, Stanford University School of Medicine, Stanford, CA 94305, USA. 2 Department of Molecular Biology, The Scripps Research Institute, La Jolla, CA 92037, USA. 3 Department of Structural Biology, Stanford University School of Medicine, Stanford, CA 94305, USA.
* These authors contributed equally to this work.
To whom correspondence should be addressed. E-mail: stevens{at}scripps.edu (R.C.S.); kobilka{at}stanford.edu (B.K.K.)
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Vadim Cherezov, Daniel M. Rosenbaum, Michael A. Hanson, Søren G. F. Rasmussen, Foon Sun Thian, Tong Sun Kobilka, Hee-Jung Choi, Peter Kuhn, William I. Weis, Brian K. Kobilka, and Raymond C. Stevens (23 November 2007) Science318 (5854), 1258.
[DOI: 10.1126/science.1150577] |Abstract »|Full Text »|PDF »|Authors' Summary »|Supporting Online Material »
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