Label-free detection of small-molecule binding to a GPCR in the membrane environment

Heym, Roland G.; Hornberger, Wilfried; Lakics, Viktor; Terstappen, Georg C.

doi:10.1016/j.bbapap.2015.04.003

Cited by 19 publications

(15 citation statements)

References 28 publications

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“…When the receptor was reconstituted in lipid nanodiscs, protein stability was enhanced and the kinetic data obtained were more similar to native receptors compared with those solubilized in detergents. Similar results were obtained for the CXCR4 receptor when the receptor was embedded in lipoparticles largely consisting of native cell membrane (Heym et al, 2015). In combination their studies demonstrate the influence of native membrane composition upon protein performance.…”

Section: Surface Plasmon Resonance Analysissupporting

confidence: 72%

Ligand Residence Time at G-protein–Coupled Receptors—Why We Should Take Our Time To Study It

et al. 2015

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Over the past decade the kinetics of ligand binding to a receptor have received increasing interest. The concept of drug-target residence time is becoming an invaluable parameter for drug optimization. It holds great promise for drug development, and its optimization is thought to reduce off-target effects. The success of long-acting drugs like tiotropium support this hypothesis. Nonetheless, we know surprisingly little about the dynamics and the molecular detail of the drug binding process. Because protein dynamics and adaptation during the binding event will change the conformation of the protein, ligand binding will not be the static process that is often described. This can cause problems because simple mathematical models often fail to adequately describe the dynamics of the binding process. In this minireview we will discuss the current situation with an emphasis on G-protein-coupled receptors. These are important membrane protein drug targets that undergo conformational changes upon agonist binding to communicate signaling information across the plasma membrane of cells.

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Section: Surface Plasmon Resonance Analysissupporting

confidence: 72%

Ligand Residence Time at G-protein–Coupled Receptors—Why We Should Take Our Time To Study It

et al. 2015

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“…Ligand‐binding assays were often performed using radio‐ and fluorophore‐labeled ligands ; however, these labeling reactions can be tedious and the labeling molecules may interrupt the interaction between ligands and receptors. Therefore, label‐free ligands have become widely used in combination with surface plasmon resonance (SPR) , quartz crystal microbalance (QCM) , surface acoustic wave , electric impedance spectroscopy (EIS) , dynamic mass redistribution optical biosensors , or impedance‐based biosensors .…”

Section: Introductionmentioning

confidence: 99%

Bio‐nanocapsule‐based scaffold improves the sensitivity and ligand‐binding capacity of mammalian receptors on the sensor chip

Iijima

Yoshimoto

Niimi

et al. 2016

Biotechnology Journal

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Mammalian receptors are recognized as target molecules for drug discovery, and chemical libraries have been screened for both potential antagonists and agonists mainly by ligand-binding assays using immobilized receptors. A bio-nanocapsule (BNC) of approximately 30 nm that displays a tandem form of the protein A-derived immunoglobulin G (IgG) Fc-binding Z domains (denoted as ZZ-BNC) has been developed for both clustering and oriented immobilization of IgGs on the solid phase of immunosensors. In this study, human IgG1 Fc-fused vascular endothelial growth factor (VEGF) receptor was immobilized through ZZ-BNC on the sensor chip of quartz crystal microbalance (ZZ-BNC-coating). When compared with direct adsorption and protein A-coating, the sensor chip showed higher sensitivity (∽46- and ∽165-fold, respectively) and larger ligand-binding capacity (∽4- and ∽18-fold, respectively). Furthermore, the number of VEGF molecules bound to its receptor increased from 0.20 (direct adsorption) to 2.06 by ZZ-BNC-coating, strongly suggesting that ZZ-BNC reduced the steric hindrance near ligand recognition sites through oriented immobilization. Similarly, the sensitivity and ligand-binding capacity of leptin and prolactin receptors were both enhanced at a level comparable to that observed for the VEGF receptor. Thus, the combination of ZZ-BNC and Fc-fused receptors could significantly improve the function of ligand-binding assays.

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“…Surface acoustic wave biosensor (SAW) sensors are a class of microelectromechanical systems (MEMS) able to measure mass differences originated by binding and unbinding of molecules to the sensor surface, along with changes in the viscoelasticity as induced by conformational rearrangements. Recently Heyn et al demonstrated the applicability of SAW for the characterization of small-molecule binding to the GPCR CXCR4 in its native conformation in the membrane environment (131) and showed complementarity to the results generated by SPR analyses. They claim that their approach obviates the need for often laborious and timeconsuming solubilization or stabilization of membrane proteins as required for SPR.…”

Section: Electromechanical Biosensorsmentioning

confidence: 89%

Binding kinetics in drug discovery ndash A current perspective

et al. 2017

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The impact of target binding kinetics (BK) on the clinical performance of therapeutic agents is presently a topic of intense debate in drug discovery. While retrospective studies suggest that BK is a differentiating parameter in marketed medicines, it is yet unclear how this information could be used to prioritize drug candidates during lead optimization. Motivated by the question whether BK can be understood and rationally optimized, we review the most relevant literature in the field, with special focus on selected examples from our organization. First we discuss structure-kinetic relationships (SKR), and how they can be influenced by factors such as conformational changes, molecular flexibility, hydrogen bonds, hydrophobicity, water molecules and (reversible-) covalent bonds. We then introduce the methodologies currently used for the investigation of BK parameters, briefly commenting on their strengths, weaknesses and future trends. Finally, we present our current perspective on the integration of BK in the drug discovery process, aiming to stimulate further thoughts on this important subject.

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Label-free detection of small-molecule binding to a GPCR in the membrane environment

Cited by 19 publications

References 28 publications

Ligand Residence Time at G-protein–Coupled Receptors—Why We Should Take Our Time To Study It

Ligand Residence Time at G-protein–Coupled Receptors—Why We Should Take Our Time To Study It

Bio‐nanocapsule‐based scaffold improves the sensitivity and ligand‐binding capacity of mammalian receptors on the sensor chip

Binding kinetics in drug discovery ndash A current perspective

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