Discovery of Novel Nonactive Site Inhibitors of the Prothrombinase Enzyme Complex

© 2016 American Chemical Society. The risk of serious bleeding is a major liability of anticoagulant drugs that are active-site competitive inhibitors targeting the Factor Xa (FXa) prothrombin (PT) binding site. The present work identifies several new classes of small molecule anticoagulants that can act as nonactive site inhibitors of the prothrombinase (PTase) complex composed of FXa and Factor Va (FVa). These new classes of anticoagulants were identified, using a novel agnostic computational approach to identify previously unrecognized binding pockets at the FXa-FVa interface. From about three million docking calculations of 281 128 compounds in a conformational ensemble of FXa heavy chains identified by molecular dynamics (MD) simulations, 97 compounds and their structural analogues were selected for experimental validation, through a series of inhibition assays. The compound selection was based on their predicted binding affinities to FXa and their ability to successfully bind to multiple protein conformations while showing selectivity for particular binding sites at the FXa/FVa interface. From these, thirty-one (31) compounds were experimentally identified as nonactive site inhibitors. Concentration-based assays further identified 10 compounds represented by four small-molecule families of inhibitors that achieve dose-independent partial inhibition of PTase activity in a nonactive site-dependent and self-limiting mechanism. Several compounds were identified for their ability to bind to protein conformations only seen during MD, highlighting the importance of accounting for protein flexibility in structure-based drug discovery approaches

Ensemble-based docking: From hit discovery to metabolism and toxicity predictions

This paper describes and illustrates the use of ensemble-based docking, i.e., using a collection of protein structures in docking calculations for hit discovery, the exploration of biochemical pathways and toxicity prediction of drug candidates. We describe the computational engineering work necessary to enable large ensemble docking campaigns on supercomputers. We show examples where ensemble-based docking has significantly increased the number and the diversity of validated drug candidates. Finally, we illustrate how ensemble-based docking can be extended beyond hit discovery and toward providing a structural basis for the prediction of metabolism and off-target binding relevant to pre-clinical and clinical trials

Interdimer zipping in the chemoreceptor signaling domain revealed by molecular dynamics simulations

Chemoreceptors are principal components of the bacterial sensory system that modulates cellular motility. They detect changes in the environment and transmit information to CheA histidine kinase, which ultimately controls cellular flagellar motors. The prototypical Tsr chemoreceptor in E. coli is a homodimer containing two principal functional modules: (i) a periplasmic ligand-binding domain and (ii) a cytoplasmic signaling domain. Chemoreceptor dimers are arranged into a trimer of dimers at the tip of the signaling domain comprising a minimal physical unit essential for enhancing the CheA activity several hundredfold. Trimers of dimers are arranged into highly ordered hexagon arrays at the cell pole; however, the mechanism underlying the trimer-of-dimer and higher order array formation remains unclear. Furthermore, molecular mechanisms of signal transduction that are likely to involve inter-dimer interactions are not fully understood. Here we apply all-atom, microsecond-time scale molecular dynamics simulations of the Tsr trimer of dimers atomic model in order to obtain further insight into potential interactions within the chemoreceptor signaling unit. We show extensive interactions between homodimers at the hairpin tip of the signaling domain, where strong hydrophobic interactions maintain binding. A subsequent zipping of homodimers is facilitated by electrostatic interactions, in particular by polar solvation energy and salt bridges that stabilize the final compact structure, which extends beyond the kinase interacting subdomain. Our study provides evidence that interdimer interactions within the chemoreceptor signaling domain are more complex than previously thought

Determining anion-quadrupole interactions among protein, DNA, and ligand molecules

Background An extensive search through the Protein Databank (about 4500 nonredundant structures) was previously completed within our lab to analyze the energetic and geometric characteristics of an understudied molecular interaction known as an anion-quadrupole (AQ) interaction. Such an interaction occurs when the positively charged edge of an aromatic ring, resulting from a quadruple moment (i.e., a dual dipole moment), renders the aromatic molecule noncovalently bound to a nearby anionic molecule. The study considered a very limited scenario of molecules that can participate in AQ interactions, consisting of the phenyl group of a phenylalanine (phe) amino acid as the aromatic participant and the carboxylate group of an aspartate (asp) or glutamate (glu) amino acid as the anionic participant. The results revealed anion-quadrupole pairs to be prevalent within most of the protein structures. It was also observed that the interaction energy for AQ pairs was heavily dependent on the angle between the anion and plane of the aromatic ring, favoring a more planar interaction. In light of these critical observations being made from such a limited scenario, only phe-glu and phe-asp pairs and in a reduced sample set of the PDB, we are now continuing this work of identifying AQ interactions using a greatly expanded strategy. We are following these four aims: 1. Optimizing the AQ-search program to run in a semi-parallel fashion and on a large cluster of processors in order to handle larger analyses, 2. Adding to our search additional anionic participants which will include non-protein structures such as DNA and small ligands, 3. Studying a subset of the AQ pairs with molecular dynamics simulations in buried and solvent exposed environments to observe non-static behavioral traits as well as the reproducibility of AQ interactions by force field parameters. 4. Building an online database for public access to our data and search program. Acknowledgments We would like to acknowledge the NSF-IGERT traineeship, Scalable Computing and Leading Edge Innovative Technologies (SCALE-IT), for providing the resources for this project

A Politics of Intellectual Property: Creating a Patent System in Revolutionary France

Economic interpretations, in particular the lens of the Industrial Revolution, have strongly influenced our understanding of the rise of intellectual property. This article examines the political origins of the 1791 patent law in France, which is usually seen as the birth of the modern patent system in that country. Although calls to reform the Old Regime's privileges of invention were increasingly frequent as the eighteenth century wore on, only the French Revolution provided the ideological resources necessary for such a transformation. The revolutionaries did more than just adopt the procedures of English legislation, such as replacing prior examination with a registration system. I argue that the new patents (brevets d'invention) reflected the Revolution's image of the ideal society-a society built on natural rights, property, and the social contract, and made of rational inventors and an enlightened public. In France, more so than in other countries, intellectual property was the child of a political revolution rather than industrial capitalism

The Order of Technological Knowledge. Crafting a New Language for Technology in France, 1750-1850

THIS DISSERTATION EXAMINES the evolution of representations of technology in France between 1750 and 1850. It proposes a history of ways of representing technology and of inscribing technical objects within texts and especially images. The periodization that I introduce starts in the mid-18th century, with the publication of the Encyclopédie and of the Description des arts et métiers, which were the first large-scale attempts to collect, codify and systematize technological knowledge in France. It ends in the mid-19th century, with the blossoming of a specific engineering culture and the triumph of a new patent system. Instead of investigating the traditional sites of technological knowledge, such as the Academy of Sciences, the École Polytechnique, and the Conservatoire des Arts et Métiers, I adopt an oblique perspective and successively look at five different ways of not only thinking about, but also of interacting with, technology: judging, classifiying, owning, rationalizing and imagining. This dissertation argues that, during this century, a new language for technology emerged, which is indicative of changing conceptions, or epistemes, of technology. Representations of technical objects moved from humanist to geometric, from realist to schematic, and from reproductive to generative. By the mid-nineteenth century, texts and images no longer represented technical objects, but directly produced them.History of Scienc

High Performance Computing Prediction of Potential Natural Product Inhibitors of SARS-CoV-2 Key Targets

This work describes using a supercomputer to perform virtual screening of natural products and natural products derivatives against several conformations of 3 proteins of SARS-CoC-2 : papain-like protease, main protease and spike protein. We analyze the common chemical features of the top molecules predicted to bind and describe the pharmacophores responsible for the predicted binding