Light Activation of Rhodopsin: Insights from Molecular Dynamics Simulations Guided by Solid-State NMR Distance Restraints
ABSTRACT Structural restraints provided by solid-state NMR measurements of the metarhodopsin II intermediate are combined with molecular dynamics simulations to help visualize structural changes in the light activation of rhodopsin. Since the timescale for the formation of the metarhodopsin II intermediate (> 1 ms) is beyond that readily accessible by molecular dynamics, we use NMR distance restraints derived from 13C dipolar recoupling measurements to guide the simulations. The simulations yield a working model for how photoisomerization of the 11-cis retinylidene chromophore bound within the interior of rhodopsin is coupled to transmembrane helix motion and receptor activation. The mechanism of activation that emerges is that multiple switches on the extracellular (or intradiscal) side of rhodopsin trigger structural changes that converge to disrupt the ionic lock between helices H3 and H6 on the intracellular side of the receptor.
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- "er isolation of various photointermediate states of rhodopsin that dis - play characteristic absorbance maxima ( Matthews et al . , 1963 ; Yoshizawa and Wald , 1964 ; Thorgeirsson et al . , 1993 ) More recent EPR studies have revised these initial estimates for conformational alterations to ϳ6 - to 10 - Å displacements ( Altenbach et al . , 2008 ; Hornak et al . , 2010 ) . Furthermore , the chemical differences between the Meta I and Meta II states simply involve the changes in protonation state , and each of these states is in equi - librium . Were large scale movements of entire helices before G protein binding involved , it is thermodynami - cally unlikely that the equilibrium between the states co"
ABSTRACT: Crucial as molecular sensors for many vital physiological processes, seven-transmembrane domain G protein-coupled receptors (GPCRs) comprise the largest family of proteins targeted by drug discovery. Together with structures of the prototypical GPCR rhodopsin, solved structures of other liganded GPCRs promise to provide insights into the structural basis of the superfamily's biochemical functions and assist in the development of new therapeutic modalities and drugs. One of the greatest technical and theoretical challenges to elucidating and exploiting structure-function relationships in these systems is the emerging concept of GPCR conformational flexibility and its cause-effect relationship for receptor-receptor and receptor-effector interactions. Such conformational changes can be subtle and triggered by relatively small binding energy effects, leading to full or partial efficacy in the activation or inactivation of the receptor system at large. Pharmacological dogma generally dictates that these changes manifest themselves through kinetic modulation of the receptor's G protein partners. Atomic resolution information derived from increasingly available receptor structures provides an entrée to the understanding of these events and practically applying it to drug design. Supported by structure-activity relationship information arising from empirical screening, a unified structural model of GPCR activation/inactivation promises to both accelerate drug discovery in this field and improve our fundamental understanding of structure-based drug design in general. This review discusses fundamental problems that persist in drug design and GPCR structural determination.Pharmacological reviews 12/2011; 63(4):901-37. DOI:10.1124/pr.110.003350 · 17.10 Impact Factor
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- "The thermodynamic stabilities of these states are not known quantitatively and will vary with receptor and ligand, although it is clear that the agonist state is very unstable (Gether et al., 1997). Intermediate and transition states between these major conformations must also exist on the activation pathway(s), which is gradually becoming clearer from a combination of structural, spectroscopic and mutational studies (Hornak et al., 2010; Tate and Schertler, 2009). Thus we are confronted with a complex conformational landscape but one which is dominated by major receptor states, which when trapped by ligands or mutations will correspond to key pharmacological phenotypes (Colquhoun, 1998). "
ABSTRACT: G protein-coupled receptors (GPCRs) are one of the most important target classes in the central nervous system (CNS) drug discovery, however the fact they are integral membrane proteins and are unstable when purified out of the cell precludes them from a wide range of structural and biophysical techniques that are used for soluble proteins. In this study we demonstrate how protein engineering methods can be used to identify mutations which can both increase the thermostability of receptors, when purified in detergent, as well as biasing the receptor towards a specific physiologically relevant conformational state. We demonstrate this method for the adenosine A(2A) receptor and muscarinic M(1) receptor. The resultant stabilised receptors (known as StaRs) have a pharmacological profile consistent with the inverse agonist conformation. The stabilised receptors can be purified in large quantities, whilst retaining correct folding, thus generating reagents suitable for a broad range of structural and biophysical studies. In the case of the A(2A)-StaR we demonstrate that surface plasmon resonance can be used to profile the association and dissociation rates of a range of antagonists, a technique that can be used to improve the in vivo efficacy of receptor antagonists.Neuropharmacology 01/2011; 60(1):36-44. DOI:10.1016/j.neuropharm.2010.07.001 · 5.11 Impact Factor
Conference Paper: Power distribution in pulse-density modulated waveforms[Show abstract] [Hide abstract]
ABSTRACT: The pulse-density modulation technique (PDM) has been used in resonant inverters because it maintains soft-switching for any output power level. Its major drawback is that subharmonics of the resonant frequency are generated. In this paper, the frequency spectra of PDM waveforms, as a function of the duty cycle and the load time constant, are investigated. The current is analyzed as an amplitude modulated waveform, and results for regular and nonregular PDM are obtained. It is shown that the nonregular PDM, with the highest number of partial sequences and the best symmetry, produces the lowest subharmonicsPower Electronics Specialists Conference, 2000. PESC 00. 2000 IEEE 31st Annual; 02/2000