New therapeutic strategies are needed to combat the tuberculosis pandemic and the spread of multidrug-resistant (MDR) and extensively drug-resistant (XDR) forms of the disease, which remain a serious public health challenge worldwide. The most urgent clinical need is to discover potent agents capable of reducing the duration of MDR and XDR tuberculosis therapy with a success rate comparable to that of current therapies for drug-susceptible tuberculosis. The last decade has seen the discovery of new agent classes for the management of tuberculosis, several of which are currently in clinical trials. However, given the high attrition rate of drug candidates during clinical development and the emergence of drug resistance, the discovery of additional clinical candidates is clearly needed. Here, we report on a promising class of imidazopyridine amide (IPA) compounds that block Mycobacterium tuberculosis growth by targeting the respiratory cytochrome bc1 complex. The optimized IPA compound Q203 inhibited the growth of MDR and XDR M. tuberculosis clinical isolates in culture broth medium in the low nanomolar range and was efficacious in a mouse model of tuberculosis at a dose less than 1 mg per kg body weight, which highlights the potency of this compound. In addition, Q203 displays pharmacokinetic and safety profiles compatible with once-daily dosing. Together, our data indicate that Q203 is a promising new clinical candidate for the treatment of tuberculosis.

Small-subunit (SSU) rRNA gene sequences were obtained by PCR from 12 Blastocystis isolates from humans, rats, and reptiles for which elongation factor 1␣ (EF-1␣) gene sequences are already available. These new sequences were analyzed by the Bayesian method in a broad phylogeny including, for the first time, all Blastocystis sequences available in the databases. Phylogenetic trees identified seven well-resolved groups plus several discrete lineages that could represent newly defined clades. Comparative analysis of SSU rRNA-and EF-1␣-based trees obtained by maximum-likelihood methods from a restricted sampling (13 isolates) revealed overall agreement between the two phylogenies. In spite of their morphological similarity, sequence divergence among Blastocystis isolates reflected considerable genetic diversity that could be correlated with the existence of potentially >12 different species within the genus. Based on this analysis and previous PCR-based genotype classification data, six of these major groups might consist of Blastocystis isolates from both humans and other animal hosts, confirming the low host specificity of Blastocystis. Our results also strongly suggest the existence of numerous zoonotic isolates with frequent animal-to-human and human-to-animal transmissions and of a large potential reservoir in animals for infections in humans.

No abstract

No abstract

Trp-Trp (WW 1 ) domains are compact modules of 38 -40 amino acids long, folded into a three-stranded ␤ sheet, and found in single or tandem repeats in over 25 unrelated cellsignaling proteins (1, 2). Although their binding to prolinebased ligands is now well described (3, 4), little is known about their precise biological function. WW domains form a new family of protein-protein interaction modules targeted to proline residues, analogous to the Src homology (SH) 3 domains (5).Based on their proline-rich sequence binding specificities, WW domains are classified into five distinct groups (6). The N-terminal WW domain of the peptidyl-prolyl isomerase Pin1, an essential regulator at mitotic entry, is grouped among class IV domains, which bind peptides containing a proline residue preceded by a phosphoserine or a phosphothreonine (pSer/ pThr-Pro motif) (7). This latter motif is found in several mitotic Pin1-binding phosphoproteins (8), including the mitotic phosphatase Cdc25 (9, 10) and the microtubule-associated tau () protein (11).Site-directed mutagenesis experiments indicate that Pin1 binds phosphoproteins through its N-terminal WW domain and that the binding site mainly implicates the conserved residues Tyr 18 and Trp 29 (7, 11), which constitute a nearly flat hydrophobic area on the molecular surface of the WW domain. WW domains interacting with the core sequence PPXY, like Yesassociated protein, use the same hydrophobic surface for molecular recognition (3,4,12). However, this hydrophobic binding site alone is not likely to explain the phospho-dependent character of the ligand binding to the Pin1 WW domain.Recently the x-ray crystal structure of the full Pin1 protein bound to a doubly phosphorylated peptide (YpSPTpSPS) from the C-terminal domain (CTD) of RNA polymerase II was reported (13). The protein-peptide interactions are essentially limited to two regions on the WW domain surface. First, a phosphate binding pocket, encompassing the side chains of Ser 11 , Arg 12 , and the backbone amide of Arg 12 , anchors the interacting phosphate moiety via several hydrogen bonds. Second, the aromatic pair Tyr 18 -Trp 29 forms a molecular clamp that constrains the proline at position ϩ1 of the interacting phosphoserine. The peptide ligand binds to the Pin1 WW domain in a N-to C-terminal orientation, in contrast to the C-to N-terminal orientation that is found for two other proline-rich peptides in complex with a group I WW domain (3,14). Other proline recognition domains, such as the SH3 domain of the Caenorhabditis elegans signaling adaptor protein Sem5, can * This project was partly executed in the framework of the Génopole de Lille. The 600-MHz NMR facility used in this study was funded by the European community, the Région Nord-Pas de Calais, CNRS, and the Institut Pasteur de Lille. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact. 1 The ab...

His-aromatic complexes, with the His located above the aromatic plane, are stabilized by π-π, δ(+)-π and/or cation-π interactions according to whether the His is neutral or protonated and the partners are in stacked or T-shape conformations. Here we attempt to probe the relative strength of these interactions as a function of the geometry and protonation state, in gas phase, in water and protein-like environments (acetone, THF and CCl4), by means of quantum chemistry calculations performed up to second order of the Møller-Plesset pertubation theory. Two sets of conformations are considered for that purpose. The first set contains 89 interactions between His and Phe, Tyr, Trp, or Ade, observed in X-ray structures of proteins and protein-ligand complexes. The second set contains model structures obtained by moving an imidazolium/imidazole moiety above a benzene ring or an adenine moiety. We found that the protonated complexes are much more stable than the neutral ones in gas phase. This higher stability is due to the electrostatic contributions, the electron correlation contributions being equally important in the two forms. Thus, π-π and δ(+)-π interactions present essentially favorable electron correlation energy terms, whereas cation-π interactions feature in addition favorable electrostatic energies. The protonated complexes remain more stable than the neutral ones in protein-like environments, but the difference is drastically reduced. Furthermore, the T-shape conformation is undoubtedly more favorable than the stacked one in gas phase. This advantage decreases in the solvents, and the stacked conformation becomes even slightly more favorable in water. The frequent occurrence of His-aromatic interactions in catalytic sites, at protein-DNA or protein-ligand interfaces and in 3D domain swapping proteins emphasize their importance in biological processes.

A molecular understanding of drug resistance mechanisms enables surveillance of the effectiveness of new antimicrobial therapies during development and deployment in the field. We used conventional drug resistance selection as well as a regime of limiting dilution at early stages of drug treatment to probe two antimalarial imidazolopiperazines, KAF156 and GNF179. The latter approach permits isolation of low-fitness mutants that might otherwise be out-competed during selection. Whole-genome sequencing of 24 independently-derived resistant P. falciparum clones revealed four parasites with mutations in the known cyclic amine resistance locus (pfcarl), and a further 20 with mutations in two previously unreported P. falciparum drug resistance genes, an acetyl-CoA transporter (pfact) and a UDP-galactose transporter (pfugt). Mutations were validated both in vitro by CRISPR editing in P. falciparum, and in vivo by evolution of resistant P. berghei mutants. Both PfACT and PfUGT were localized to the endoplasmic reticulum by fluorescence microscopy. As mutations in pfact and pfugt conveyed resistance against additional unrelated chemical scaffolds, these genes are likely to be involved in broad mechanisms of antimalarial drug resistance.

No abstract