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- Chen, Yue, et al.
(författare)
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Allosteric Effect of Nanobody Binding on Ligand-Specific Active States of the beta 2 Adrenergic Receptor
- 2021
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Ingår i: Journal of Chemical Information and Modeling. - : American Chemical Society (ACS). - 1549-9596 .- 1549-960X. ; 61:12, s. 6024-6037
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Tidskriftsartikel (refereegranskat)abstract
- Nanobody binding stabilizes G-protein-coupled receptors (GPCR) in a fully active state and modulates their affinity for bound ligands. However, the atomic-level basis for this allosteric regulation remains elusive. Here, we investigate the conformational changes induced by the binding of a nanobody (Nb80) on the active-like beta 2 adrenergic receptor (beta 2AR) via enhanced sampling molecular dynamics simulations. Dimensionality reduction analysis shows that Nb80 stabilizes structural features of the beta 2AR with an similar to 14 angstrom outward movement of transmembrane helix 6 and a close proximity of transmembrane (TM) helices 5 and 7, and favors the fully active-like conformation of the receptor, independent of ligand binding, in contrast to the conditions under which no intracellular binding partner is bound, in which case the receptor is only stabilized in an intermediateactive state. This activation is supported by the residues located at hotspots located on TMs 5, 6, and 7, as shown by supervised machine learning methods. Besides, ligand-specific subtle differences in the conformations assumed by intracellular loop 2 and extracellular loop 2 are captured from the trajectories of various ligand-bound receptors in the presence of Nb80. Dynamic network analysis further reveals that Nb80 binding triggers tighter and stronger local communication networks between the Nb80 and the ligand-binding sites, primarily involving residues around ICL2 and the intracellular end of TM3, TM5, TM6, as well as ECL2, ECL3, and the extracellular ends of TM6 and TM7. In particular, we identify unique allosteric signal transmission mechanisms between the Nb80-binding site and the extracellular domains in conformations modulated by a full agonist, BI167107, and a G-protein-biased partial agonist, salmeterol, involving mainly TM1 and TM2, and TM5, respectively. Altogether, our results provide insights into the effect of intracellular binding partners on the GPCR activation mechanism, which should be taken into account in structure-based drug discovery.
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| 2. |
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| 3. |
- Fleetwood, Oliver, 1990-, et al.
(författare)
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Identification of ligand-specific G protein-coupled receptor states and prediction of downstream efficacy via data-driven modeling
- 2021
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Ingår i: eLIFE. - : eLife Sciences Publications, Ltd. - 2050-084X. ; 10
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Tidskriftsartikel (refereegranskat)abstract
- Ligand binding stabilizes different G protein-coupled receptor states via a complex allosteric process that is not completely understood. Here, we have derived free energy landscapes describing activation of the beta(2) adrenergic receptor bound to ligands with different efficacy profiles using enhanced sampling molecular dynamics simulations. These reveal shifts toward active-like states at the Gprotein-binding site for receptors bound to partial and full agonists, and that the ligands modulate the conformational ensemble of the receptor by tuning protein microswitches. We indeed find an excellent correlation between the conformation of the microswitches close to the ligand binding site and in the transmembrane region and experimentally reported cyclic adenosine monophosphate signaling responses. Dimensionality reduction further reveals the similarity between the unique conformational states induced by different ligands, and examining the output of classifiers highlights two distant hotspots governing agonism on transmembrane helices 5 and 7.
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| 4. |
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| 5. |
- Fleetwood, Oliver, 1990-
(författare)
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New approaches to data-driven analysis and enhanced sampling simulations of G protein-coupled receptors
- 2021
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Proteins are large biomolecules that carry out specific functions within living organisms. Understanding how proteins function is a massive scientific challenge with a wide area of applications. In particular, by controlling protein function we may develop therapies for many diseases. To understand a protein’s function, we need to consider its full conformational ensemble, and not only a single snapshot of a structure. Allosteric signaling is a factor often driving protein conformation change, where the binding of a molecule to one site triggers a response in another part of the protein. G protein-coupled receptors (GPCRs) are transmembrane proteins that bind molecules outside the membrane, which enables coupling to a G protein in their intracellular domain. Understanding the complex allosteric process governing this mechanism could have a significant impact on the development of novel drugs.Molecular dynamics (MD) is a computational method that can capture protein conformational change at an atomistic level. However, MD is a computationally expensive approach to simulating proteins, and is thus infeasible for many applications. Enhanced sampling techniques have emerged to reduce the computational cost of standard MD. Another challenge with MD is to extract useful information and distinguish signal from noise in an MD trajectory. Data-driven methods can streamline analysis of protein simulations and improve our understanding of biomolecular systems.Paper 1 and 2 contain methodological developments to analyze the results of MD in a data-driven manner. We provide methods that create interpretable maps of important molecular features from protein simulations (Paper 1) and identify allosteric communication pathways in biological systems (Paper 2). As a result, more insights can be extracted from MD trajectories. Our approach is generalizable and can become useful to analyze complex simulations of various biomolecular systems. In Paper 3 and 4, we combine the aforementioned methodological advancements with enhanced sampling techniques to study a prototypical GPCR, the β2 adrenergic receptor. First, we make improvements to the string method with swarms of trajectories and derive the conformational change and free energy along the receptor’s activation pathway. Next, we identify key molecular microswitches directly or allosterically controlled by orthosteric ligands and show how these couple to a shift in probability of the receptor’s active state. In Paper 4, we also find that ligands induce ligand-specific states, and the molecular basis governing these states. These new approaches generate insights compatible with previous simulation and experimental studies at a relatively low computational cost. Our work also provides new insights into the molecular basis of allosteric communication in membrane proteins, and might become a useful tool in the design of novel GPCR drugs.
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