Genetically Encoded Photo-cross-linkers Map the Binding Site of an Allosteric Drug on a G Protein-Coupled Receptor

G protein-coupled receptors (GPCRs) are dynamic membrane proteins that bind extracellular molecules to transduce signals. Although GPCRs represent the largest class of therapeutic targets, only a small percentage of their ligand-binding sites are precisely defined. Here we describe the novel application of targeted photo-cross-linking using unnatural amino acids to obtain structural information about the allosteric binding site of a small molecule drug, the CCR5-targeted HIV-1 co-receptor blocker maraviroc

Controlling the charge transfer flow at the graphene/pyrene-nitrilotriacetic acid interface

The fabrication of highly efficient bio-organic nanoelectronic devices is still a challenge due to the difficulty in interfacing the biomolecular component to the organic counterparts. One of the ways to overcome this bottleneck is to add a self-assembled monolayer (SAM) in between the electrode and the biological material. The addition of a pyrene-nitrilotriacetic acid layer to a graphene metal electrode enhances the charge transfer within the device. Our theoretical calculations and electrochemical results show that the formation of a pyrene-nitrilotriacetic acid SAM enforces a direct electron transfer from graphene to the SAM, while the addition of the Ni2+ cation and imidazole reverses the charge transfer direction, allowing an atomic control of the electron flow, which is essential for a true working device. © 2018 The Royal Society of Chemistry

Ligand- and mutation-induced conformational selection in the CCR5 chemokine G protein-coupled receptor

We predicted the structural basis for pleiotropic signaling of the C-C chemokine type 5 (CCR5) G protein-coupled receptor (GPCR) by predicting the binding of several ligands to the lower-energy conformations of the CCR5 receptor and 11 mutants. For each case, we predicted the ∼20 most stable conformations for the receptor along with the binding sites for four anti-HIV ligands. We found that none of the ligands bind to the lowest-energy apo-receptor conformation. The three ligands with a similar pharmacophore (Maraviroc, PF-232798, and Aplaviroc) bind to a specific higher-energy receptor conformation whereas TAK-779 (with a different pharmacophore) binds to a different high-energy conformation. This result is in agreement with the very different binding-site profiles for these ligands obtained by us and others. The predicted Maraviroc binding site agrees with the recent structure of CCR5 receptor cocrystallized with Maraviroc. We performed 11 site-directed mutagenesis experiments to validate the predicted binding sites. Here, we independently predicted the lowest 10 mutant protein conformations for each of the 11 mutants and then docked the ligands to these lowest conformations. We found the predicted binding energies to be in excellent agreement with our mutagenesis experiments. These results show that, for GPCRs, each ligand can stabilize a different protein conformation, complicating the use of cocrystallized structures for ligand screening. Moreover, these results show that a single-point mutation in a GPCR can dramatically alter the available low-energy conformations, which in turn alters the binding site, potentially altering downstream signaling events. These studies validate the conformational selection paradigm for the pleiotropic function and structural plasticity of GPCRs

Finding class C GPCR subtype-discriminating n-grams through feature selection

G protein-coupled receptors (GPCRs) are a large and heterogeneous superfamily of receptors that are key cell players for their role as extracellular signal transmitters. Class C GPCRs, in particular, are of great interest in pharmacology. The lack of knowledge about their full 3-D structure prompts the use of their primary amino acid sequences for the construction of robust classifiers, capable of discriminating their different subtypes. In this paper, we describe the use of feature selection techniques to build Support Vector Machine (SVM)-based classification models from selected receptor subsequences described as n-grams. We show that this approach to classification is useful for finding class C GPCR subtype-specific motifs.Peer ReviewedPostprint (author’s final draft

From the Committee of 100 to the Committee to Re-Elect the President: The Political Campaigns of Richard M. Nixon

From the Committee of 100 to the Committee to Re-elect the President: The Political Campaigns of Richard M. Nixon offers the reader a comprehensive biography of Richard M. Nixon through the lens of his political campaigns. This thesis illustrates how Richard Nixon became one of the fiercest campaigners in 20th century American political history. This thesis, furthermore, examines the key staff and strategy of each campaign Nixon waged. This thesis, additionally, presents to the reader insight on how Nixon often fought his campaigns independently from the Republican Party and how he relied on the help of a few dedicated men

Manipulation by design

Human behaviour is affected by architecture, including how online user interfaces are designed. The purpose of this article is to provide insights into the regulation of behaviour modification by the design of choice architecture in light of the European Union data protection law (GDPR) and marketing law (UCPD). It has become popular to use the term ‘dark pattern’ (also ‘deceptive practices’) to describe such practices in online environments. The term provides a framework for identifying and discussing ‘problematic’ design practices, but the definitions and descriptions are not sufficient in themselves to draw the fine line between legitimate (lawful) persuasion and unlawful manipulation, which requires an inquiry into agency, self-determination, regulation and legal interpretation. The main contribution of this article is to place manipulative design, including ‘dark patterns’, within the framework of persuasion (marketing), technology (persuasive technology) and law (privacy and marketing).</p

Active Learning and its Impact on Higher-Order Thinking Skills in Preschool Science Education

Active learning strategies engage preschool children to use higher order thinking skills such as critical thinking and problem solving, however this method of teaching is underused in favor of traditional methods.  A background and literature review will establish the effectiveness of using active learning strategies with early learners in science instruction.  This article will address reasons teachers do not implement higher order learning strategies in the classroom and attempt to refute these reasons.  This article establishes that a teacher does not need to have “mastered” this strategy in order to implement it.  Suggestions and examples will be given for how to transform a standard preschool lesson into one that engages higher order thinking skills through the use of active learning strategies

Common Genetic Variants Explain the Majority of the Correlation Between Height and Intelligence : The Generation Scotland Study

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Use of G-Protein-Coupled and -Uncoupled CCR5 Receptors by CCR5 Inhibitor-Resistant and -Sensitive Human Immunodeficiency Virus Type 1 Variants

Small-molecule CCR5 inhibitors such as vicriviroc (VVC) and maraviroc (MVC) are allosteric modulators that impair HIV-1 entry by stabilizing a CCR5 conformation that the virus recognizes inefficiently. Viruses resistant to these compounds are able to bind the inhibitor-CCR5 complex while also interacting with the free coreceptor. CCR5 also interacts intracellularly with G proteins, as part of its signal transduction functions, and this process alters its conformation. Here we investigated whether the action of VVC against inhibitor-sensitive and -resistant viruses is affected by whether or not CCR5 is coupled to G proteins such as Gαi. Treating CD4^(+)T cells with pertussis toxin to uncouple the Gαi subunit from CCR5 increased the potency of VVC against the sensitive viruses and revealed that VVC-resistant viruses use the inhibitor-bound form of Gα_(i)-coupled CCR5 more efficiently than they use uncoupled CCR5. Supportive evidence was obtained by expressing a signaling-deficient CCR5 mutant with an impaired ability to bind to G proteins, as well as two constitutively active mutants that activate G proteins in the absence of external stimuli. The implication of these various studies is that the association of intracellular domains of CCR5 with the signaling machinery affects the conformation of the external and transmembrane domains and how they interact with small-molecule inhibitors of HIV-1 entry