Molecular Basis of Ligand Dissociation in β-Adrenergic Receptors

Abstract

The important and diverse biological functions of β-adrenergic receptors (βARs) have promoted the search for compounds to stimulate or inhibit their activity. In this regard, unraveling the molecular basis of ligand binding/unbinding events is essential to understand the pharmacological properties of these G protein-coupled receptors. In this study, we use the steered molecular dynamics simulation method to describe, in atomic detail, the unbinding process of two inverse agonists, which have been recently co-crystallized with β1 and β2ARs subtypes, along four different channels. Our results indicate that this type of compounds likely accesses the orthosteric binding site of βARs from the extracellular water environment. Importantly, reconstruction of forces and energies from the simulations of the dissociation process suggests, for the first time, the presence of secondary binding sites located in the extracellular loops 2 and 3 and transmembrane helix 7, where ligands are transiently retained by electrostatic and Van der Waals interactions. Comparison of the residues that form these new transient allosteric binding sites in both βARs subtypes reveals the importance of non-conserved electrostatic interactions as well as conserved aromatic contacts in the early steps of the binding process.

Effect of channel mutations on the uptake and release of the retinal ligand in opsin

“…After light-induced formation of Meta II, the photolyzed agonist all-trans-retinal leaves the protein through the open Ops* conformation, which then returns to the Ops conformation. Although other GPCRs do not bind their ligand covalently, the uptake of ligands and their positioning in the binding pocket may obey similar rules as described here for rhodopsin (39).…”

GPCRs through the keyhole: the role of protein flexibility in ligand binding to β-adrenoceptors

“…We also know that there is a great deal of conformational flexibility possible in the area of the keyhole associated with receptor activation -2-AR structures show a 2.1 Å movement of the Cα of Ser 5.46 when agonist and antagonist structures are compared (a difference of ~1 Å in the 1-AR turkey structures (González et al, 2011;Warne et al, 2011)). We also observe various rotamer states of Ser 5.46 depending on whether it can directly H-bond to the ligand occupying the binding site, so our hypothesis that a keyhole can form in this region seems plausible.…”

Beyond Standard Molecular Dynamics: Investigating the Molecular Mechanisms of G Protein-Coupled Receptors with Enhanced Molecular Dynamics Methods

“…While multiple long timescale atomistic standard MD simulations (of the order of hundreds of µs) have recently permitted researchers to visualize, for instance, the binding of different ligands to the B2AR (Dror et al 2011), biased MD techniques have been successfully employed to enhance the probability of observing such events during shorter simulation timescales. Three studies we discuss in this section focus on the premise that from knowledge of the pathways by which a ligand can exit a receptor, one can infer specific details of the binding mechanism (Gonzalez et al 2011; Selvam et al 2012; Wang and Duan 2009). The probability of spontaneous ligand exit from a binding cavity, on the timescale accessible to MD simulations, can be enhanced by addition of an external force imposed upon the ligand, as is the case for steered MD and random acceleration MD.…”