Cosolvent-Based Protein Pharmacophore for Ligand Enrichment in Virtual Screening.

Virtual screening of large compound databases, looking for potential ligands of a target protein, is a major tool in computer-aided drug discovery. Throughout the years, different techniques such as similarity searching, pharmacophore matching, or molecular docking have been applied with the aim of finding hit compounds showing appreciable affinity. Molecular dynamics simulations in mixed solvents have been shown to identify hot spots relevant for protein-drug interaction, and implementations based on this knowledge were developed to improve pharmacophore matching of small molecules, binding free-energy estimations, and docking performance in terms of pose prediction. Here, we proved in a retrospective manner that cosolvent-derived pharmacophores from molecular dynamics (solvent sites) improve the performance of docking-based virtual screening campaigns. We applied a biased docking scheme based on solvent sites to nine relevant target proteins that have a set of known ligands or actives and compounds that are, presumably, nonbinders (decoys). Our results show improvement in virtual screening performance compared to traditional docking programs both at a global level, with up to 35% increase in areas under the receiver operating characteristic curve, and in early stages, with up to a 7-fold increase in enrichment factors at 1%. However, the improvement in pose prediction of actives was less profound. The presented application makes use of the AutoDock Bias method and is the only cosolvent-derived pharmacophore technique that employs its knowledge both in the ligand conformational search algorithm and the final affinity scoring for virtual screening purposes.

HNO Is Produced by the Reaction of NO with Thiols.

Azanone (nitroxyl, HNO) is a highly reactive compound whose biological role is still a matter of debate. One possible route for its formation is NO reduction by biological reductants. These reactions have been historically discarded due to the negative redox potential for the NO,H+/HNO couple. However, the NO to HNO conversion mediated by vitamins C, E, and aromatic alcohols has been recently shown to be feasible from a chemical standpoint. Based on these precedents, we decided to study the reaction of NO with thiols as potential sources of HNO. Using two complementary approaches, trapping by a Mn porphyrin and an HNO electrochemical sensor, we found that under anaerobic conditions aliphatic and aromatic thiols (as well as selenols) are able to convert NO to HNO, albeit at different rates. Further mechanistic analysis using ab initio methods shows that the reaction between NO and the thiol produces a free radical adduct RSNOH•, which reacts with a second NO molecule to produce HNO and a nitrosothiol. The nitrosothiol intermediate reacts further with RSH to produce a second molecule of HNO and RSSR, as previously reported.

LigQ: A Webserver to Select and Prepare Ligands for Virtual Screening.

Virtual screening is a powerful methodology to search for new small molecule inhibitors against a desired molecular target. Usually, it involves evaluating thousands of compounds (derived from large databases) in order to select a set of potential binders that will be tested in the wet-lab. The number of tested compounds is directly proportional to the cost, and thus, the best possible set of ligands is the one with the highest number of true binders, for the smallest possible compound set size. Therefore, methods that are able to trim down large universal data sets enriching them in potential binders are highly appreciated. Here we present LigQ, a free webserver that is able to (i) determine best structure and ligand binding pocket for a desired protein, (ii) find known binders, as well as potential ligands known to bind to similar protein domains, (iii) most importantly, select a small set of commercial compounds enriched in potential binders, and (iv) prepare them for virtual screening. LigQ was tested with several proteins, showing an impressive capacity to retrieve true ligands from large data sets, achieving enrichment factors of over 10%. LigQ is available at http://ligq.qb.fcen.uba.ar/ .

Germline hypomorphic CARD11 mutations in severe atopic disease.

Few monogenic causes for severe manifestations of common allergic diseases have been identified. Through next-generation sequencing on a cohort of patients with severe atopic dermatitis with and without comorbid infections, we found eight individuals, from four families, with novel heterozygous mutations in CARD11, which encodes a scaffolding protein involved in lymphocyte receptor signaling. Disease improved over time in most patients. Transfection of mutant CARD11 expression constructs into T cell lines demonstrated both loss-of-function and dominant-interfering activity upon antigen receptor-induced activation of nuclear factor-κB and mammalian target of rapamycin complex 1 (mTORC1). Patient T cells had similar defects, as well as low production of the cytokine interferon-γ (IFN-γ). The mTORC1 and IFN-γ production defects were partially rescued by supplementation with glutamine, which requires CARD11 for import into T cells. Our findings indicate that a single hypomorphic mutation in CARD11 can cause potentially correctable cellular defects that lead to atopic dermatitis.