Structure-activity relationships of actively FhuE transported rifabutin derivatives with potent activity against Acinetobacter baumannii.

Hospital-acquired infections are on the rise and represent both, a clinical and financial burden. With resistance emerging and an ever-dwindling armamentarium at hand, infections caused by Acinetobacter baumannii are particularly problematic, since these bacteria have a high level of resistance and resilience to traditional and even last-resort antibiotics. The antibiotic rifabutin was recently found to show potent in vitro and in vivo activity against extensively drug resistant A. baumannii. Building on this discovery, we report on the synthesis and activity of rifabutin analogs, with a focus on N-functionalization of the piperidine ring. The antimicrobial testing uncovered structure activity relationships (SAR) for A. baumannii that were not reflected in Staphylococcus aureus. The cellular activity did not correlate with cell-free transcription inhibition, but with bacterial intracellular compound accumulation. Mass spectrometry-based accumulation studies confirmed the involvement of the siderophore receptor FhuE in active compound translocation at low concentrations, and they showed a strong impact of the culture medium on the accumulation of rifabutin. Overall, the study underlines the structural feature required for strong accumulation of rifabutin in A. baumannii and identifies analogs as or more potent than rifabutin against A. baumannii.

Discovery pipelines for marine resources: an ocean of opportunity for biotechnology?

Marine microbial diversity offers enormous potential for discovery of compounds of crucial importance in healthcare, food security and bioindustry. However, access to it has been hampered by the difficulty of accessing and growing the organisms for study. The discovery and exploitation of marine bioproducts for research and commercial development requires state-of-the-art technologies and innovative approaches. Technologies and approaches are advancing rapidly and keeping pace is expensive and time consuming. There is a pressing need for clear guidance that will allow researchers to operate in a way that enables the optimal return on their efforts whilst being fully compliant with the current regulatory framework. One major initiative launched to achieve this, has been the advent of European Research Infrastructures. Research Infrastructures (RI) and associated centres of excellence currently build harmonized multidisciplinary workflows that support academic and private sector users. The European Marine Biological Research Infrastructure Cluster (EMBRIC) has brought together six such RIs in a European project to promote the blue bio-economy. The overarching objective is to develop coherent chains of high-quality services for access to biological, analytical and data resources providing improvements in the throughput and efficiency of workflows for discovery of novel marine products. In order to test the efficiency of this prototype pipeline for discovery, 248 rarely-grown organisms were isolated and analysed, some extracts demonstrated interesting biochemical properties and are currently undergoing further analysis. EMBRIC has established an overarching and operational structure to facilitate the integration of the multidisciplinary value chains of services to access such resources whilst enabling critical mass to focus on problem resolution.

Biophysical characterization of E. coli TolC interaction with the known blocker hexaamminecobalt.

BACKGROUND: The tripartite efflux pump AcrAB-TolC in E. coli is involved in drug resistance by transporting antibiotics out of the cell. The outer membrane protein TolC can be blocked by various cations, including hexaamminecobalt, thereby TolC represents a potential target for reducing antimicrobial resistance as its blockage may improve efficacy of antibiotics. METHODS: We utilized single channel electrophysiology measurements for studying TolC conductance in the absence and presence of the known TolC blocker hexaamminecobalt. Association and dissociation constants of hexaamminecobalt were determined using surface plasmon resonance measurements. Minimum inhibitory concentration (MIC) assays in the absence and presence of antibiotics were carried out for investigating the antibacterial effect of hexaamminecobalt and its potential to reduce MICs. RESULTS: TolC gating in the absence of any ligand is voltage dependent and asymmetric at high applied voltages. Hexaamminecobalt binds to TolC with high affinity and kinetic data revealed fast association and dissociation rates. Despite potent binding to TolC, hexaamminecobalt does not possess an intrinsic antimicrobial activity against E. coli nor does it reduce MIC values of antibiotics erythromycin and fusidic acid. CONCLUSIONS: TolC opening can be effectively blocked by small molecules. More potent channel blockers are needed in order to investigate the eligibility of TolC as drug target. GENERAL SIGNIFICANCE: TolC, a potentially interesting pharmaceutical target can be addressed by small molecules, blocking the channel. Biophysical characterization of the binding processes will support future identification and optimisation of more potent TolC blockers in order to validate TolC as a pharmaceutical target.

Bifunctional antimicrobial conjugates and hybrid antimicrobials.

Covering: up to the end of 2016Novel antimicrobial drugs are continuously needed to counteract bacterial resistance development. An innovative molecular design strategy for novel antibiotic drugs is based on the hybridization of an antibiotic with a second functional entity. Such conjugates can be grouped into two major categories. In the first category (antimicrobial hybrids), both functional elements of the hybrid exert antimicrobial activity. Due to the dual targeting, resistance development can be significantly impaired, the pharmacokinetic properties can be superior compared to combination therapies with the single antibiotics, and the antibacterial potency is often enhanced in a synergistic manner. In the second category (antimicrobial conjugates), one functional moiety controls the accumulation of the other part of the conjugate, e.g. by mediating an active transport into the bacterial cell or blocking the efflux. This approach is mostly applied to translocate compounds across the cell envelope of Gram-negative bacteria through membrane-embedded transporters (e.g. siderophore transporters) that provide nutrition and signalling compounds to the cell. Such 'Trojan Horse' approaches can expand the antibacterial activity of compounds against Gram-negative pathogens, or offer new options for natural products that could not be developed as standalone antibiotics, e.g. due to their toxicity.

Target identification by image analysis.

Covering: 1997 to the end of 2015Each biologically active compound induces phenotypic changes in target cells that are characteristic for its mode of action. These phenotypic alterations can be directly observed under the microscope or made visible by labelling structural elements or selected proteins of the cells with dyes. A comparison of the cellular phenotype induced by a compound of interest with the phenotypes of reference compounds with known cellular targets allows predicting its mode of action. While this approach has been successfully applied to the characterization of natural products based on a visual inspection of images, recent studies used automated microscopy and analysis software to increase speed and to reduce subjective interpretation. In this review, we give a general outline of the workflow for manual and automated image analysis, and we highlight natural products whose bacterial and eucaryotic targets could be identified through such approaches.

Antiviral drug discovery: broad-spectrum drugs from nature.

Covering: up to April 2014. The development of drugs with broad-spectrum antiviral activities is a long pursued goal in drug discovery. It has been shown that blocking co-opted host-factors abrogates the replication of many viruses, yet the development of such host-targeting drugs has been met with scepticism mainly due to toxicity issues and poor translation to in vivo models. With the advent of new and more powerful screening assays and prediction tools, the idea of a drug that can efficiently treat a wide range of viral infections by blocking specific host functions has re-bloomed. Here we critically review the state-of-the-art in broad-spectrum antiviral drug discovery. We discuss putative targets and treatment strategies, with particular focus on natural products as promising starting points for antiviral lead development.

Platinum dioxide cation: easy to generate experimentally but difficult to describe theoretically.

A formal platinum(V) dioxide cation [Pt,O2](+) can be generated in the gas phase by successive oxidation of Pt(+) with N2O. The ion's reactivity is in keeping with the dioxide structure OPtO(+), rather than with [Pt,O2](+) isomers having intact O-O bonds, e.g., the dioxygen complex Pt(O2)(+) and peroxo species PtOO(+). Inter alia due to the high ionization energy of the neutral counterpart (11.2 eV), the [Pt,O2](+) cation is a rather aggressive reagent toward oxidizable neutrals. [Pt,O2](+) is even capable of activating inert substrates such as H2, CO, and CH4. Further, a sequence for the catalytic conversion CO + N(2)O --> CO2 + N2 is described with a turnover number of >100 for the catalytically active species PtOn(+) (n = 0-2). As a consequence of the high reactivity, however, the observed selectivities with most substrates are rather poor. For example, the reaction of PtO2(+) with ethane gives rise to 10 different product channels. In an attempt to analyze the structural features and different minima of the [Pt,O2](+) system, extensive ab initio studies are performed. While correlated ab initio methods describe the system reasonably well, density functional theory turns out to be much less accurate in terms of both structural and energetic descriptions.

A mechanistic study of the FeO+-mediated decomposition pathways of phenol, anisol, and their thio analogues.

The gas-phase oxidations of phenol, anisol, thiophenol, and thioanisol by 'bare' FeO+ are examined by using Fourier transform-ion cyclotron resonance (FT-ICR) and tandem mass-spectrometry. Reaction mechanisms are derived on the basis of isotope-labeling experiments, MS/MS studies, and comparison with structural isomers, that is ions formed by independent routes. The chemistry of all substrates is determined by the functional groups, whereas reactions typical of unsubstituted benzene with FeO+ are suppressed. For phenol and thiophenol, four-membered metallacycles are obtained concomitant with a regioselective loss of water, which involves the O atom from the FeO+ entity and hydrogen atoms originating from the functional group and from the ortho position of the ring. C-H bond cleavage of the methoxy group (kH/kD = 2.0) is rate-contributing for the degradation of metastable anisol/FeO+, which is featured by highly regioselective losses of H2O, HCO, H2CO, and [C,H2,O2]. In the oxidation of thioanisol, two different C-H bond activation mechanisms are operating, resulting in the elimination of [Fe,H,O,S] concomitant with the formation of the benzyl cation (kH/kD = 4.7), and loss of water (kH/kD = 2.5). The reactions of independently generated, formal S- and C-oxidation intermediates of thioanisol indicate the occurrence of extensive structural isomerizations prior to dissociation. For anisol and thioanisol, analogies and differences between oxidation reactions catalyzed by the enzyme cytochrome P-450 in the condensed phase and those observed for the gas-phase model FeO+ are discussed.