Using knowledge of the substrate specificity of cPLA(2) (phospholipases A(2)), a novel series of inhibitors of this enzyme were designed based upon a three point model of inhibitor binding to the enzyme active site comprising a lipophilic anchor, an electrophilic serine "trap", and an acidic binding moiety. The resulting 1,3-diheteroatom-substituted propan-2-ones were evaluated as inhibitors of cPLA(2) in both aggregated bilayer and soluble substrate assays. Systematic variation of the lipophilic, electrophilic, and acidic groups revealed a well-defined structure-activity relationship against the enzyme. Optimization of each group led to compound 22 (AR-C70484XX), which contains a decyloxy lipophilic side chain, a 1,3-diaryloxypropan-2-one moiety as a unique serine trap, and a benzoic acid as the acidic binding group. AR-C70484XX was found to be among the most potent in vitro inhibitors of cPLA(2) described to date being more than 20-fold more active against the isolated enzyme (IC(50) = 0.03 microM) than the standard cPLA(2) inhibitor, arachidonyl trifluoromethyl ketone (AACOCF(3)), and also greater than 10-fold more active than AACOCF(3) against the cellular production of arachidonic acid by HL60 cells (IC(50) = 2.8 microM).

The Staphylococcus aureus multidrug-binding protein QacR represses transcription of the plasmid-encoded membrane protein QacA, a multidrug efflux transporter. QacR is induced by multiple structurally dissimilar monovalent and bivalent cationic lipophilic compounds, many of which are effluxed from the cell by QacA via the proton motive force. The multidrug-binding pocket of QacR has been shown to be quite extensive and features several glutamates and multiple aromatic residues. To date, the structure of only one QacR-bivalent cationic drug complex (that of QacR bound to dequalinium) has been determined, and how other longer or shorter bivalent cationic compounds bind is unknown. Here we report the crystal structures of QacR bound to two cytotoxic bivalent diamidines, pentamidine and hexamidine. These compounds are structurally similar, differing by only one methylene carbon in the alkyl chain linker. However, this small difference results in very dissimilar binding modes. Similar to dequalinium, hexamidine spans the multidrug-binding pocket, and its positively charged benzamidine groups are neutralized by residues Glu-57 and Glu-120. Pentamidine binds QacR in a novel fashion whereby one of its benzamidine groups interacts with residue Glu-63, and the other is neutralized by carbonyl and side chain oxygen atoms. Thus, these structures demonstrate that a formal negative charge is not a prerequisite for binding positively charged drugs and underscore the versatility of the QacR and, likely, all multidrug-binding pockets.

The approval of bedaquiline to treat tuberculosis has validated adenosine triphosphate (ATP) synthase as an attractive target to kill Mycobacterium tuberculosis (Mtb). Herein, we report the discovery of two diverse lead series imidazo[1,2-a]pyridine ethers (IPE) and squaramides (SQA) as inhibitors of mycobacterial ATP synthesis. Through medicinal chemistry exploration, we established a robust structure-activity relationship of these two scaffolds, resulting in nanomolar potencies in an ATP synthesis inhibition assay. A biochemical deconvolution cascade suggested cytochrome c oxidase as the potential target of IPE class of molecules, whereas characterization of spontaneous resistant mutants of SQAs unambiguously identified ATP synthase as its molecular target. Absence of cross resistance against bedaquiline resistant mutants suggested a different binding site for SQAs on ATP synthase. Furthermore, SQAs were found to be noncytotoxic and demonstrated efficacy in a mouse model of tuberculosis infection.

The aim of this 15-month feeding study was to investigate the effects of more 16 sustainable feeds on specific growth rate, fatty acid composition and 17 metabolism of Arctic charr (Salvelinus alpinus). A control feed, formulated with 18 fish meal and fish oil (F1), was compared with feeds where the marine 19 ingredients were increasingly replaced by pumpkin kernel cake and rapeseed oil 20 (Feeds F2, F3, and F4). Arctic charr were randomly distributed into 12 tanks and 21 fed one of the feeds in triplicate. The biomass of fish fed F1 and F2 diets was 22 significantly higher compared to fish fed diet F4 with highest replacement level. 23However, the dorsal and ventral muscle tissues had very similar total saturated, 24 monounsaturated, and polyunsaturated fatty acid (PUFA) contents, irrespective 25 of dietary supply. Although diets F3 and F4 contained 6-fold less fish oil than 26 diets F1 and F2, fish fed diets F3 and F4 retained only 2-fold less highly desired 27 omega-3 (n-3) long-chain (LC)-PUFA in their dorsal and ventral muscle tissues. 28Incubating isolated hepatocytes with 14 C-labeled α-linolenic acid (18:3n-3) 29 provided evidence that Arctic charr can bioconvert this essential dietary PUFA to 30 n-3 LC-PUFA, including docosahexaenoic acid. The results suggested that tissue 31 fatty acid compositions in Arctic charr are dependent, not only on dietary fatty 32 acid supply, but also on their ability for endogenous synthesis of n-3 LC-PUFA. 33Finally, this long-term feeding study indicated that feeds containing 34 pumpkinseed press cake and rapeseed oil produced fish with largely similar fatty 35 acid composition to fish fed diets containing higher contents of fish meal and 36 fish oil. 37 38