Christopher A. Reynolds

The magnetic nature of disk accretion onto black holes

Although disk accretion onto compact objects—white dwarfs, neutron stars and black holes—is central to much of high-energy astrophysics, the mechanisms that enable this process have remained observationally difficult to determine. Accretion disks must transfer angular momentum in order for matter to travel radially inward onto the compact object. Internal viscosity from magnetic processes and disk winds can both in principle transfer angular momentum, but hitherto we lacked evidence that either occurs. Here we report that an X-ray-absorbing wind discovered in an observation of the stellar-mass black hole binary GRO J1655 - 40 (ref. 6) must be powered by a magnetic process that can also drive accretion through the disk. Detailed spectral analysis and modelling of the wind shows that it can only be powered by pressure generated by magnetic viscosity internal to the disk or magnetocentrifugal forces. This result demonstrates that disk accretion onto black holes is a fundamentally magnetic process.

Dimerization and Domain Swapping in G-Protein-Coupled Receptors: A Computational Study

In recent years there has been an increasing number of reports describing G protein-coupled receptor (GPCR) dimerization and heterodimerization. However, the evidence on the nature of the dimers and their role in GPCR activation is inconclusive. Consequently, we present here a review of our computational studies on G protein-coupled receptor dimerization and domain swapping. The studies described include molecular dynamics simulations on receptor monomers and dimers in the absence of ligand, in the presence of an agonist, and in the presence of an antagonist (or more precisely an inverse agonist). Two distinct sequence-based approaches to studying protein interfaces are also described, namely correlated mutation analysis and evolutionary trace analysis. All three approaches concur in supporting the proposal that the dimerization interface includes transmembrane helices 5 and 6. These studies cannot distinguish between domain swapped dimers and contact dimers as the models used were restricted to the helical part of the receptor. However, it is proposed that for the purpose of signalling, the domain swapped dimer and the corresponding contact dimer are equivalent. The evolutionary trace analysis suggests that every GPCR family and subfamily (for which sufficient sequence data is available) has the potential to dimerize through this common functional site on helices 5 and 6. The evolutionary trace results on the G protein are briefly described and these are consistent with GPCR dimerization. In addition to the functional site on helices 5 and 6, the evolutionary trace analysis identified a second functional site on helices 2 and 3. Possible roles for this site are suggested, including oligomerization.

Dimerization of G-protein-coupled receptors.

The evolutionary trace (ET) method, a data mining approach for determining significant levels of amino acid conservation, has been applied to over 700 aligned G-protein-coupled receptor (GPCR) sequences. The method predicted the occurrence of functionally important clusters of residues on the external faces of helices 5 and 6 for each family or subfamily of receptors; similar clusters were observed on helices 2 and 3. The probability that these clusters are not random was determined using Monte Carlo techniques. The cluster on helices 5 and 6 is consistent with both 5,6-contact and 5,6-domain swapped dimer formation; the possible equivalence of these two types of dimer is discussed because this relates to activation by homo- and heterodimers. The observation of a functionally important cluster of residues on helices 2 and 3 is novel, and some possible interpretations are given, including heterodimerization and oligomerization. The application of the evolutionary trace method to 113 aligned G-protein sequences resulted in the identification of two functional sites. One large, well-defined site is clearly identified with adenyl cyclase, beta/gamma and regulator of G-protein signaling (RGS) binding. The other G-protein functional site, which extends from the ras-like domain onto the helical domain, has the correct size and electrostatic properties for GPCR dimer binding. The implications of these results are discussed in terms of the conformational changes required in the G-protein for activation by a receptor dimer. Further, the implications of GPCR dimerization for medicinal chemistry are discussed in the context of these ET results.

Towards new transition metal-based hypoxic selective agents for therapy and imaging.

The greater lability of Co(II) relative to Co(III) can potentially be used to achieve selective delivery of nitrogen mustard type molecules to hypoxic cells. Attempts to improve the stability of the Co(II) state by utilising tripodal tetradentate ligands are described, together with the results of DF calculations. Rhenium has two beta-emitting isotopes (186)Re and (188)Re that have potential for use to treat cancer if the complexes can be targeted with sufficient specificity. We describe some new rapid low temperature routes using hydrazines to labile Re(V) and Re(III) species which provide potential convenient access to a wide range of oxo- and diazenido-complexes. The synthesis of new Re(V) and Re(III) thiosemicarbazone complexes is presented in the context of obtaining hypoxic selective species. Copper(II) bis(thiosemicarbazone) complexes are known to be hypoxic selective and spectroscopic, cyclic voltammetric and computational studies of the mechanism are presented, together with the synthesis of new Cu(II) complexes directed towards the hypoxic selective delivery of nitrogen mustard type molecules.

Toward the active conformations of rhodopsin and the β2‐adrenergic receptor

Using sets of experimental distance restraints, which characterize active or inactive receptor conformations, and the X‐ray crystal structure of the inactive form of bovine rhodopsin as a starting point, we have constructed models of both the active and inactive forms of rhodopsin and the β2‐adrenergic G‐protein coupled receptors (GPCRs). The distance restraints were obtained from published data for site‐directed crosslinking, engineered zinc binding, site‐directed spin‐labeling, IR spectroscopy, and cysteine accessibility studies conducted on class A GPCRs. Molecular dynamics simulations in the presence of either “active” or “inactive” restraints were used to generate two distinguishable receptor models. The process for generating the inactive and active models was validated by the hit rates, yields, and enrichment factors determined for the selection of antagonists in the inactive model and for the selection of agonists in the active model from a set of nonadrenergic GPCR drug‐like ligands in a virtual screen using ligand docking software. The simulation results provide new insights into the relationships observed between selected biochemical data, the crystal structure of rhodopsin, and the structural rearrangements that occur during activation. Proteins 2004.

Free energy calculations in molecular biophysics

The free energy perturbation method, and the related thermodynamic integration methods, have recently had a great influence on theoretical chemists interested in applying computational techniques to study problems of biological importance. The infinite order free energy perturbation equation was derived almost forty years ago, but only within the last five years has it been widely used in conjunction with Monte Carlo or molecular dynamics simulations to calculate free energy differences between similar macromolecular systems in solution. The purpose of this review is firstly to describe the formalism behind the methods but secondly and more importantly to describe the wide range of problems to which it has been applied. Here the focus is primarily on problems of chemical or biological interest rather than on problems studied for the sake of methodological development. The methods, of necessity, are generally used in conjunction with empirical force fields and the limitations arising from this and related ...

Bioinformatics and molecular modelling approaches to GPCR oligomerization

The elusive nature of the structure and function of the G-protein coupled receptor (GPCR) dimer or oligomer has led to a variety of computational studies, most of which have been directed primarily towards understanding structure. Here we review some of the recent studies based on sequence analysis and docking experiments and the recent developments in GPCR structure that have underpinned dimerization studies. In addition, we review recent nanosecond molecular dynamics simulations and coarse-grained methods for investigating the dynamic consequences of dimerization. The strengths and weaknesses of these complementary methods are discussed. The consensus of a variety of studies is that several transmembrane helices are involved in the dimerization/oligomerization interface(s); computation has been particularly effective in elucidating the experiments that seem to indicate a key role for transmembrane helix 4.

The Extracellular Surface of the GLP-1 Receptor Is a Molecular Trigger for Biased Agonism

Summary Ligand-directed signal bias offers opportunities for sculpting molecular events, with the promise of better, safer therapeutics. Critical to the exploitation of signal bias is an understanding of the molecular events coupling ligand binding to intracellular signaling. Activation of class B G protein-coupled receptors is driven by interaction of the peptide N terminus with the receptor core. To understand how this drives signaling, we have used advanced analytical methods that enable separation of effects on pathway-specific signaling from those that modify agonist affinity and mapped the functional consequence of receptor modification onto three-dimensional models of a receptor-ligand complex. This yields molecular insights into the initiation of receptor activation and the mechanistic basis for biased agonism. Our data reveal that peptide agonists can engage different elements of the receptor extracellular face to achieve effector coupling and biased signaling providing a foundation for rational design of biased agonists.

The oxidation potential of 1,4-diaminobenzene: Calculation versus experiment

Abstract The oxidation potential of 1,4-diaminobenzene has been calculated theoretically using a combination of ab-initio calculations and molecular dynamics simulations. An experimental determination of the same quantity is presented. Data obtained from cyclic voltammetric experiments, at a platinum electrode, were consistent with an EC electrode reaction mechanism. Cyclic voltammograms recorded at high voltage scan rates ( v ⪢ 500 mV s −1 ), such that the following homogeneous kinetics were outrun, indicated the oxidation to be quasi-reversible and this enabled the standard oxidation potential to be estimated. Comparison of the theoretical and experimental values for the oxidation potential showed agreement to within 25 mV. The level of agreement between theory and experiment was considered to be highly satisfactory.

Errors in free-energy perturbation calculations due to neglecting the conformational variation of atomic charges

Abstract There is much concern about the accuracy of free-energy perturbation calculations. Here we have investigated the problem of ignoring the conformational variation of atomic charges by calculating the difference in the free energy of hydration between ethanol and propanol and comparing the results obtained for conformationally transferable charges with those obtained using conventional atomic charges and experiment. The results show that the common practice of ignoring the conformational variation of atomic charges may lead to serious errors which can dominate over all other errors.