Biophysical Mapping of the Adenosine A2A Receptor

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• 作者：Andrei Zhukov ; Stephen P. Andrews ; James C. Errey ; Nathan Robertson ; Benjamin Tehan ; Jonathan S. Mason ; Fiona H. Marshall ; Malcolm Weir ; Miles Congreve
• 刊名：Journal of Medicinal Chemistry
• 出版年：2011
• 出版时间：July 14, 2011
• 年：2011
• 卷：54
• 期：13
• 页码：4312-4323
• 全文大小：1168K
• 年卷期：v.54,no.13(July 14, 2011)
• ISSN：1520-4804

文摘

A new approach to generating information on ligand receptor interactions within the binding pocket of G protein-coupled receptors has been developed, called Biophysical Mapping (BPM). Starting from a stabilized receptor (StaR), minimally engineered for thermostability, additional single mutations are then added at positions that could be involved in small molecule interactions. The StaR and a panel of binding site mutants are captured onto Biacore chips to enable characterization of the binding of small molecule ligands using surface plasmon resonance (SPR) measurement. A matrix of binding data for a set of ligands versus each active site mutation is then generated, providing specific affinity and kinetic information (K_D, k_on, and k_off) of receptor鈥搇igand interactions. This data set, in combination with molecular modeling and docking, is used to map the small molecule binding site for each class of compounds. Taken together, the many constraints provided by these data identify key protein鈥搇igand interactions and allow the shape of the site to be refined to produce a high quality three-dimensional picture of ligand binding, thereby facilitating structure based drug design. Results of biophysical mapping of the adenosine A_2A receptor are presented.