Biacore analysis with stabilized G-protein-coupled receptors

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• 作者：Rebecca L. Rich ; James Errey ; Fiona Marshall ; David G. Myszka
• 关键词：Surface plasmon resonance (SPR) ; Protein– ; protein ; Kinetics
• 刊名：Analytical Biochemistry
• 出版年：2011
• 出版时间：15 February 2011
• 年：2011
• 卷：409
• 期：2
• 页码：267-272
• 全文大小：1138 K

文摘

Using stabilized forms of β₁ adrenergic and A_2A adenosine G-protein-coupled receptors, we applied Biacore to monitor receptor activity and characterize binding constants of small-molecule antagonists spanning more than 20,000-fold in affinity. We also illustrate an improved method for tethering His-tagged receptors on NTA (carboxymethylated dextran preimmobilized with nitrilotriacetic acid) chips to yield stable, high-capacity, high-activity surfaces as well as a novel approach to regenerate receptor binding sites. Based on our success with this approach, we expect that the combination of stabilized receptors with biosensor technology will become a common method for characterizing members of this receptor family.