Search

Rapid advancements in information processing and embedded systems require high selective and fast sensors. Conventional gas sensors are not suitable for the detection of isomers of organic compounds due to cross-sensitivity and the response time being limited by slow chemical kinetics. Amperometric gas sensors using conducting polymers modified with metal catalysts are a suitable and robust system due to many tunable properties. In this paper, conducting polymer polyaniline was electrochemically decorated with clusters containing precisely defined number of gold atoms to function as an electro-catalyst. The modified polymer composite showed fast reaction rate for the electro-oxidation of alcohols in both liquid and gas phases. The number of gold atoms affected the catalytic activity. Cyclic voltammograms were measured and results showed discriminable patterns between n-propanol and iso-propanol even at different gas concentrations. Thus, it was demonstrated that gas sensor arrays can be realized by decorating different number of gold atoms on polyaniline electrodes, to yield defined and different selectivity.

Antitumor pyrrolobenzodiazepines (PBDs), lincosamide antibiotics, quorum-sensing molecule hormaomycin, and antimicrobial griselimycin are structurally and functionally diverse groups of actinobacterial metabolites. The common feature of these compounds is the incorporation of l-tyrosine- or l-leucine-derived 4-alkyl-l-proline derivatives (APDs) in their structures. Here, we report that the last reaction in the biosynthetic pathway of APDs, catalyzed by F420H2-dependent Apd6 reductases, contributes to the structural diversity of APD precursors. Specifically, the heterologous overproduction of six Apd6 enzymes demonstrated that Apd6 from the biosynthesis of PBDs and hormaomycin can reduce only an endocyclic imine double bond, whereas Apd6 LmbY and partially GriH from the biosyntheses of lincomycin and griselimycin, respectively, also reduce the more inert exocyclic double bond of the same 4-substituted Δ1-pyrroline-2-carboxylic acid substrate, making LmbY and GriH unusual, if not unique, among reductases. Furthermore, the differences in the reaction specificity of the Apd6 reductases determine the formation of the fully saturated APD moiety of lincomycin versus the unsaturated APD moiety of PBDs, providing molecules with optimal shapes to bind their distinct biological targets. Moreover, the Apd6 reductases establish the first F420H2-dependent enzymes from the luciferase-like hydride transferase protein superfamily in the biosynthesis of bioactive molecules. Finally, our bioinformatics analysis demonstrates that Apd6 and their homologues, widely distributed within several bacterial phyla, play a role in the formation of novel yet unknown natural products with incorporated l-proline-like precursors and likely in the microbial central metabolism.

In natural environments, antibiotics are important means of interspecies competition. At subinhibitory concentrations, they act as cues or signals inducing antibiotic production; however, our knowledge of well-documented antibiotic-based sensing systems is limited. Here, for the soil actinobacterium Streptomyces lincolnensis, we describe a fundamentally new ribosome-mediated signaling cascade that accelerates the onset of lincomycin production in response to an external ribosome-targeting antibiotic to synchronize antibiotic production within the population. The entire cascade is encoded in the lincomycin biosynthetic gene cluster (BGC) and consists of three lincomycin resistance proteins in addition to the transcriptional regulator LmbU: a lincomycin transporter (LmrA), a 23S rRNA methyltransferase (LmrB), both of which confer high resistance, and an ATP-binding cassette family F (ABCF) ATPase, LmrC, which confers only moderate resistance but is essential for antibiotic-induced signal transduction. Specifically, antibiotic sensing occurs via ribosome-mediated attenuation, which activates LmrC production in response to lincosamide, streptogramin A, or pleuromutilin antibiotics. Then, ATPase activity of the ribosome-associated LmrC triggers the transcription of lmbU and consequently the expression of lincomycin BGC. Finally, the production of LmrC is downregulated by LmrA and LmrB, which reduces the amount of ribosome-bound antibiotic and thus fine-tunes the cascade. We propose that analogous ABCF-mediated signaling systems are relatively common because many ribosome-targeting antibiotic BGCs encode an ABCF protein accompanied by additional resistance protein(s) and transcriptional regulators. Moreover, we revealed that three of the eight coproduced ABCF proteins of S. lincolnensis are clindamycin responsive, suggesting that the ABCF-mediated antibiotic signaling may be a widely utilized tool for chemical communication. IMPORTANCE Resistance proteins are perceived as mechanisms protecting bacteria from the inhibitory effect of their produced antibiotics or antibiotics from competitors. Here, we report that antibiotic resistance proteins regulate lincomycin biosynthesis in response to subinhibitory concentrations of antibiotics. In particular, we show the dual character of the ABCF ATPase LmrC, which confers antibiotic resistance and simultaneously transduces a signal from ribosome-bound antibiotics to gene expression, where the 5' untranslated sequence upstream of its encoding gene functions as a primary antibiotic sensor. ABCF-mediated antibiotic signaling can in principle function not only in the induction of antibiotic biosynthesis but also in selective gene expression in response to any small molecules targeting the 50S ribosomal subunit, including clinically important antibiotics, to mediate intercellular antibiotic signaling and stress response induction. Moreover, the resistance-regulatory function of LmrC presented here for the first time unifies functionally inconsistent ABCF family members involving antibiotic resistance proteins and translational regulators.

Cerium oxide nanoparticles (CeNPs) possess multiple redox enzyme mimetic activities in scavenging reactive oxygen species (ROS) as a potential biomedicine. These enzymatic activities of CeNPs are closely related to their surface oxidation state. Here we have reported a synthetic method to modify CeNPs’ surface oxidation state by changing the conformation of the poly(acrylic acid) (PAA) polymers adsorbed onto the CeNP surface. The synthesized PAA–CeNPs exhibited the same core size, morphology, crystal structure, and colloidal stability, with the only variation being their surface oxidation state (Ce3+ percentage). The modification mechanism can be attributed to the polymers chemisorbed onto the metal oxide surface forming a metal complexation structure. Such adsorption further modified CeNPs’ surface oxidation state in a temperature-dependent manner. The series of PAA–CeNPs exhibited multiple redox enzyme mimetic activities (superoxide dismutase, catalase, peroxidase, and oxidase) directly related to their surface oxidation state. In vitro experiments showed no cytotoxic effect of these PAA–CeNPs on the osteoblastic cell line SAOS-2 at high loadings. Microscopic images confirmed the internalization of PAA–CeNPs in the cells. All tested PAA–CeNPs can reduce the basal and hydrogen peroxide-induced intracellular ROS level in the cells, indicating their effective intracellular ROS scavenging effect. However, we did not observe a positive correlation between the CeNP surface oxidation state and their capacities to reduce the intracellular ROS levels. We propose that CeNPs can maintain a dynamic state of Ce3+/Ce4+ during their catalytic activities, exhibiting a non-linear correlation between the CeNP surface oxidation state and their effect on regulating the intracellular ROS level.

The modified layout of the barrier method called liquid chromatography under limiting conditions of enthalpic interactions is presented. It enables automated quantitative separation of blends of synthetic polymers, for example the single step discrimination of both parent homopolymers from the block copolymers. Moreover, this method enables the estimation of molar mass and molar mass distribution of the block copolymer precursor. Adjacent large sequences of mobile phase of different composition are applied as barriers. They are created by a computer controlled pair of pumping systems in the form of longitudinal profiles along the column. The home synthesized block copolymers polystyrene-block-poly(2-vinylpyridine) served as model examples of the method application. The adsorption retention mechanism was exploited using mesoporous bare silica gel column packing. Series of block copolymers of similar composition can be quickly handled with the method to optimize their synthesis.

The interaction between a double-hydrophilic comb copolymer with the polyanionic backbone poly[methacrylic acid-stat-poly(ethylene glycol) methyl ether methacrylate] (PMAA–PEGMA) and the cationic s...

Graphene oxide (GO) has high potential for grafting of polymer chains. This mostly enhances exfoliation or controls structure-directing effects in thermoplastic blends, whereas various complex arrays may be formed in the case of epoxy thermosets. This study reveals that the structure of these arrays is markedly influenced by parameters of grafted polymer chains and geometry of grafting. The best-balanced properties were found for nacre-mimicking lamellar arrays with an important role of internal structure and elasticity of grafted chains. Beneficial is s relatively thick “elastic interlayer” of grafted butadiene‐acrylonitrile copolymer with toughening effect comparable to much higher content of free polymer addition.

In natural environments, antibiotics are an important instrument of inter-species competition. At subinhibitory concentrations, they act as cues or signals inducing antibiotic production: however, our knowledge of well-documented antibiotic-based sensing systems is limited. Here, for the soil actinobacterium Streptomyces lincolnensis we describe a fundamentally new ribosome-mediated signaling cascade that accelerates the onset of lincomycin production in response to an external ribosome-targeting antibiotic to synchronize the antibiotic production within the population. The entire cascade is encoded within the lincomycin biosynthetic gene cluster (BGC) and besides the transcriptional regulator, LmbU it consists of three lincomycin resistance proteins: a lincomycin transporter, LmrA, a 23S rRNA methyltransferase, LmrB, both conferring a high resistance, and an ABCF ATPase LmrC that confers only moderate resistance but is indispensable for the antibiotic-induced signal transduction. Specifically, the antibiotic sensing occurs via a ribosome-mediated attenuation, which activates LmrC production in response to lincosamide, streptogramin A, or pleuromutilin antibiotics. Then, the ribosome-operating LmrC ATPase activity triggers the transcription of lmbU and consequently the expression of lincomycin BGC. Finally, the production of LmrC is downregulated by LmrA and LmrB which reduces the amount of the ribosome-bound antibiotic and thus fine-tune the cascade. We propose that analogous ABCF-mediated signaling systems are relatively common because many BGCs for ribosome-targeting antibiotics encode an ABCF-protein accompanied by additional resistance protein(s) and transcriptional regulators. Moreover, we revealed that three of eight co-produced ABCF proteins of S. lincolnensis are clindamycin-responsive thus the ABCF-mediated antibiotic signaling might be generally utilized tool of chemical communication.IMPORTANCEResistance proteins are perceived as mechanisms protecting bacteria from the inhibitory effect of their produced antibiotic or antibiotics from competitors. Here, we report that antibiotic resistance proteins regulate lincomycin biosynthesis in response to subinhibitory concentrations of antibiotics. Particularly, we show the dual character of ABCF ATPase LmrC which confers antibiotic resistance and simultaneously transduces a signal from ribosome-bound antibiotic to gene expression, where the 5’ untranslated sequence upstream of its encoding gene functions as a primary antibiotic sensor. The ABCF-mediated antibiotic signaling can in principle function not only in the induction of antibiotic biosynthesis but in general in selective gene expression in response to any small molecules targeting the 50S ribosomal subunit, including clinically important antibiotics, to mediate intercellular antibiotic signaling and stress response induction. Moreover, the resistance-regulatory function of LmrC presented here for the first time unifies yet functionally inconsistent ABCF family involving the antibiotic resistance proteins and the translational regulators.

Heme oxygenase (HO) and biliverdin reductase (BVR) activities are important for neuronal function and redox homeostasis. Resuscitation from cardiac arrest (CA) frequently results in neuronal injury and delayed neurodegeneration that typically affect vulnerable brain regions, primarily hippocampus (Hc) and motor cortex (mC), but occasionally also striatum and cerebellum. We questioned whether these delayed effects are associated with changes of the HO/BVR system. We therefore analyzed the activities of HO and BVR in the brain regions Hc, mC, striatum and cerebellum of rats subjected to ventricular fibrillation CA (6 min or 8 min) after 2 weeks following resuscitation, or sham operation. From all investigated regions, only Hc and mC showed significantly decreased HO activities, while BVR activity was not affected. In order to find an explanation for the changed HO activity, we analyzed protein abundance and mRNA expression levels of HO-1, the inducible, and HO-2, the constitutively expressed isoform, in the affected regions. In both regions we found a tendency for a decreased immunoreactivity of HO-2 using immunoblots and immunohistochemistry. Additionally, we investigated the histological appearance and the expression of markers indicative for activation of microglia [tumor necrosis factor receptor type I (TNFR1) mRNA and immunoreactivity for ionized calcium-binding adapter molecule 1 (Iba1])], and activation of astrocytes [immunoreactivity for glial fibrillary acidic protein (GFAP)] in Hc and mC. Morphological changes were detected only in Hc displaying loss of neurons in the cornu ammonis 1 (CA1) region, which was most pronounced in the 8 min CA group. In this region also markers indicating inflammation and activation of pro-death pathways (expression of HO-1 and TNFR1 mRNA, as well as Iba1 and GFAP immunoreactivity) were upregulated. Since HO products are relevant for maintaining neuronal function, our data suggest that neurodegenerative processes following CA may be associated with a decreased capacity to convert heme into HO products in particularly vulnerable brain regions.

In recent experiments, the “local pH” near polyelectrolyte chains was determined from the shift in the effective acidity constant of fluorescent pH indicators attached to the macromolecules. This indirect determination raises the question if the analyzed quantity was indeed the “local pH” and what this term actually means. In this study, we combined experiments and simulations to demonstrate that the shift in ionization constant is slightly lower than the difference between the pH and the “local pH”. This offset is caused by correlations between fluctuations in chain conformation, small-ion distribution, and fluorophore ionization.

Natural pyrrolobenzodiazepines (PBDs) form a large and structurally diverse group of antitumour microbial metabolites produced through complex pathways, which are encoded within biosynthetic gene clusters. We sequenced the gene cluster of limazepines and proposed their biosynthetic pathway based on comparison with five available gene clusters for the biosynthesis of other PBDs. Furthermore, we tested two recombinant proteins from limazepine biosynthesis, Lim5 and Lim6, with the expected substrates in vitro. The reactions monitored by LC-MS revealed that limazepine biosynthesis involves a new way of 3-hydroxyanthranilic acid formation, which we refer to as the chorismate/DHHA pathway and which represents an alternative to the kynurenine pathway employed for the formation of the same precursor in the biosynthesis of other PBDs. The chorismate/DHHA pathway is presumably also involved in the biosynthesis of PBD tilivalline, several natural products unrelated to PBDs, and its part is shared also with phenazine biosynthesis. The similarities between limazepine and phenazine biosynthesis indicate tight evolutionary links between these groups of compounds.

The modification of epoxy with polymer-grafted graphene oxide (GO) is a radical alternative to mere combination of GO with a polymeric modifier. Here, this aspect is revealed for the case of bimodal grafting of GO with hydrophobic polybutadiene and hydrophilic polyoxypropylene or polyethylene glycol utilizing grafting-to approach to link different chains in the plane and edge area of GO. The results indicate fair ability of bimodally grafted GO to form ordered lamellar structures in epoxy. However, these structures show larger distance between less aligned GO plates in contrast to tightly aligned GO arrangement in case of single grafting. This is enabled by repulsive interactions between dissimilar chains. Such a structure with higher mobility of grafted chains has favorable effect on mechanical performance of epoxy. The effect of chain type and geometry of grafting is highlighted as well. Important finding is promotion of lamellar structure formation also by grafted low-m. w. compounds.

Aims: Rapid restoration of sinus rhythm is an integral part of the management of recent-onset atrial fibrillation. We aimed to assess safety and efficacy of vernakalant, a multi-channel blocking agent, in combination with external electrical cardioversion. Methods: This prospective cohort study comprised 230 patients (female 35%; median age 50 IQR 42–55) with recent-onset AF presenting to a university tertiary care center during a 6-year period. Management included intravenous vernakalant followed by electrical cardioversion in case of pharmacological failure. Results: Within 11 min (IQR 8–29), sinus rhythm could be restored by sole pharmacological management in 167 patients (73%). A left ventricular function lower than 55% (OR 3.51 (1.45–8.52)) and prior atrial fibrillation episodes being classified as persistent (OR 2.33 (1.13–4.80)) were significant predictors for non-response to vernakalant. Electrical cardioversion was successful in all patients but one within 196 min (IQR 149–300) of administration of first dosage of vernakalant. No serious adverse events could be observed. 3 patients needed further in-patient care. Conclusion: Management of recent-onset atrial fibrillation consisting of intravenous vernakalant followed by electrical cardioversion in case of failure appears safe and efficacious. Achieving a rapid conversion, this approach could potentially save resources and costs. Keywords: Vernakalant, Recent-onset atrial fibrillation, Cardioversion

Dalbavancin, a lipoglycopeptide with prolonged half-life approved for the treatment of acute bacterial skin and soft tissue infections, can be used for the treatment of infections caused by gram-positive bacteria requiring long term treatment such as endocarditis, prosthetic joint infections (PJI) or osteomyelitis. Clinical data are limited in these settings. Objectives: To evaluate indications, safety, tolerability and long-term outcomes of dalbavancin-treated patients.Patients and methods Our multicenter, retrospective study includes patients who received dalbavancin in Austria from September 2016 to March 2018. 90-day outcomes and tolerability were determined. Results: A total of 101 patients were included in 3 centers (57% male, median age 65 years). The treated infections were PJI (31%), osteomyelitis (29%), endocarditis (25%) and acute bacterial skin and soft tissue infections (12%). Concomitant use of other antimicrobial substances was common (63%). The mean total cumulative dose of dalbavancin was 3,357 mg (±2,283 mg). Clinical success rate was 89%. Side effects occurred in 3/101 patients. Conclusion: In this real-life study dalbavancin was primarily used in off-label indications for treatment of PJI, osteomyelitis and endocarditis. Success rate was high (89%), tolerability and safety were excellent in this setting. Dalbavancin may therefore be used in these off-label indications as alternative treatment approach. Keywords: Dalbavancin, Osteomyelitis, PJI, Endocarditis

New sulfonamides 5/6 derived from 4-methoxyacetophenone 1 were synthesized by N-sulfonation reaction of ammonia (3) and aminopyrimidinone (4) with its sulfonyl chloride derivative 2. Sulfonamides 5 and 6 were used as precursors of two new series of chalcones 8a-f and 9a-f, which were obtained through Claisen-Schmidt condensation with aromatic aldehydes 7a-f. Compounds 5/6, 8a-d, 8f, 9a-d, and 9f were screened by the US National Cancer Institute (NCI) at 10 μM against sixty different human cancer cell lines (one-dose trial). Chalcones 8b and 9b satisfied the pre-determined threshold inhibition criteria and were selected for screening at five different concentrations (100, 10, 1.0, 0.1, and 0.01 μM). Compound 8b exhibited remarkable GI50 values ranging from 0.57 to 12.4 μM, with cytotoxic effects being observed in almost all cases, especially against the cell lines K-562 of Leukemia and LOX IMVI of Melanoma with GI50 = 0.57 and 1.28 μM, respectively. Moreover, all compounds were screened against Mycobacterium tuberculosis H37Rv, chalcones 8a-c and 9a-c were the most active showing MIC values between 14 and 42 μM, and interestingly they were devoid of antibacterial activity against Mycobacterium smegmatis and Staphylococcus aureus. These antituberculosis hits showed however low selectivity, being equally inhibitory to M. tuberculosis and mammalian T3T cells. The chalcone-sulfonamide hybrids 8a-f and 9a-f resulted to be appealing cytotoxic agents with significant antituberculosis activity.

The immediate post-condensation steps in lincomycin biosynthesis are reminiscent of the mycothiol-dependent detoxification system of actinomycetes. This machinery provides the last proven lincomycin intermediate, a mercapturic acid derivative, which formally represents the ‘waste product’ of the detoxification process. We identified and purified new lincomycin intermediates from the culture broth of deletion mutant strains of Streptomyces lincolnensis and tested these compounds as substrates for proteins putatively involved in lincomycin biosynthesis. The results, based on LC-MS, in-source collision-induced dissociation mass spectrometry and NMR analysis, revealed the final steps of lincomycin biosynthesis, i.e. conversion of the mercapturic acid derivative to lincomycin. Most importantly, we show that deacetylation of the N′-acetyl-S-cysteine residue of the mercapturic acid derivative is required to ‘escape’ the detoxification-like system and proceed towards completion of the biosynthetic pathway. Additionally, our results, supported by L-cysteine-13C3, 15N incorporation experiments, give evidence that a different type of reaction catalysed by the homologous pair of pyridoxal-5′-phosphate-dependent enzymes, LmbF and CcbF, forms the branch point in the biosynthesis of lincomycin and celesticetin, two related lincosamides.

Formation of interpolyelectrolyte complexes (IPECs) of poly(methacrylic acid) (PMAA) bearing a fluorescent label (umbelliferone) at the chain end and poly[3,5-bis(trimethyl ammoniummethyl)-4-hydroxystyrene iodide]-block-poly(ethylene oxide) (QNPHOS-PEO) acting as a fluorescence quencher, was followed using a combination of scattering, calorimetry, microscopy and fluorescence spectroscopy techniques. While scattering and microscopy measurements indicated formation of spherical core/corona nanoparticles with the core of the QNPHOS/PMAA complex and the PEO corona, fluorescence measurements showed that both static and dynamic quenching efficiency were increased in the nanoparticle stability region. As the dynamic quenching rate constant remained unchanged, the quenching enhancement was caused by the increase in the local concentration of QNPHOS segments in the microenvironment of the label. This finding implies that the local dynamics of PMAA end chains affecting the interaction of the label with QNPHOS segments was independent of both PMAA and QNPHOS chain conformations.

Covering: up to 2017 This review covers the biosynthetic and evolutionary aspects of lincosamide antibiotics, antitumour pyrrolobenzodiazepines (PBDs) and the quorum-sensing molecule hormaomycin. These structurally and functionally diverse groups of complex natural products all incorporate rarely occurring 4-alkyl-L-proline derivatives (APDs) biosynthesized from L-tyrosine through an unusual specialized pathway catalysed by a common set of six proteins named Apd1–Apd6. We give an overview of APD formation, which involves unusual enzyme activities, and its incorporation, which is based either on nonribosomal peptide synthetase (PBDs, hormaomycin) or a unique hybrid ergothioneine-dependent condensation system followed by mycothiol-dependent sulphur atom incorporation (lincosamides). Furthermore, within the public databases, we identified 36 novel unannotated biosynthetic gene clusters that putatively encode the biosynthesis of APD compounds. Their products presumably include novel PBDs, but also novel classes of APD compounds, indicating an unprecedented potential for the diversity enhancement of these functionally versatile complex metabolites. In addition, phylogenetic analysis of known and novel gene clusters for the biosynthesis of APD compounds allowed us to infer novel evolutionary hypotheses: Apd3 methyltransferase originates from a duplication event in a hormaomycin biosynthetic gene cluster ancestor, while putative Apd5 isomerase is evolutionarily linked to PhzF protein from the biosynthesis of phenazines. Lastly, we summarize the achievements in preparing hybrid APD compounds by directing their biosynthesis, and we propose that the number of nature-like APD compounds could by multiplied by replacing L-proline residues in various groups of complex metabolites with APD, i.e. by imitating the natural process that occurs with lincosamides and PBDs, in which the replacement of L-proline for APD has proved to be an evolutionary successful concept.

The p(MMA-b-t-BuA) copolymer was prepared using the initiation system methyl 2-lithioisobutyrate/lithium/tert-butoxide (MIB-Li/t-BuOLi). A low-molecular-weight fraction was isolated and analyzed by MALDI-TOF mass spectrometry with the aim to find out structures of products of self-termination, which proceeds mainly after the addition of t-BuA into the living PMMA. The main self-termination products are formed after addition of one or two t-BuA units onto living PMMA-Li by backbiting reaction of t-BuA-Li chain-ends and carbonyl group of the antepenultimate MMA units of the same chain. The oxoesters generated in this way contain hydroxygroup, and its hydrogen can protonize any of the living chains, regardless of the type of its terminal unit. Signals of the chains, terminated by this protonization, have also been found in the spectra. Thus, the first direct evidences for the structures of the self-termination products in the anionic methacrylate/acrylate block copolymerization were obtained in this way.

In this study, we determined the Pd(II) chlorocomplex species that has the most favorable interaction with an electropolymerized and protonated polyaniline (PANI) film. This study was completed with the intent to use this species to electrochemically build atomic palladium clusters in the PANI matrix. Varying amounts of NaCl were added to a K2PdCl4/HClO4 solution to result in three species studied: PdCl2(H2O)2, PdCl3(H2O)−, and PdCl42–. UV–vis spectroscopy was used to confirm the speciation, and Raman spectroscopy, X-ray photoelectron spectroscopy, and cyclic voltammograms were used to probe the interaction between the Pd species and PANI. It was determined that PdCl3(H2O)− most effectively interacts with PANI as a result of the charge balance between the anion and the protonated nitrogen-containing groups in the polymer. It has been also found that some fraction of inserted Pd(II) cannot be reduced to Pd(0).

For the first time, ATRP was successfully employed for homopolymerization of a commercial methacrylate-functionalized polyhedral oligomeric silsesquioxane (POSS) monomer, iBuPOSSMA, to high molecular weights. It was found that iBuPOSSMA has a low ceiling temperature (Tc); therefore, low temperatures and/or high initial monomer concentrations need to be used in order to avoid low degrees of polymerization that had been observed previously. The values of Tc, as well as of the polymerization enthalpy ΔHp and entropy ΔSp were determined to be 130 °C (at [M]0 = 1 M), −41 kJ mol–1, and −101 J mol–1 K–1, respectively. Under optimized conditions, poly(iBuPOSSMA) homopolymers having low dispersity and high Mn, ranging from 23 000 to 460 000, were obtained in a well-controlled ATRP process. Moreover, various block copolymers having high-Mn poly(iBuPOSSMA) blocks were prepared by copolymerization of iBuPOSSMA with methyl methacrylate and styrene.

A series of P(MMA- co -GMA)- b -PMAPOSS block copolymers (BCPs) were synthesized by ATRP. The BCPs contain POSS (polyhedral oligomeric silsesquioxane) as a pendant unit on a particular polymer block chain. In selective solvents, the BCPs self-assemble to form ordered micellar-like structures. Spherical, cylindrical or vesicle-like morphologies were produced by tuning the BCP and mixed solvent compositions. Crosslinking of the BCP by the reaction of the glycidyl groups in the P(MMA- co -GMA) block of the micelle shell (GMA = glycidyl methacrylate) with a diamine results in a long-range structure ordering. The hexagonally packed cylindrical arrangement of the BCP networks was revealed by SAXS and TEM. The hierarchical, ordered structure of the POSS-containing hybrids was fixed by crosslinking. In addition, transient physical gels were prepared from triblock copolymers with outer MAPOSS blocks, and their rheological behavior was investigated.

Chemical diversity: Two SAM-dependent N-methyltransferases-LmbJ from the biosynthesis of the antibiotic lincomycin and CcbJ from celesticetin biosynthesis-have been characterized and compared. Both tested enzymes form multimers and are able to utilize N-demethyllincomycin, the natural substrate of LmbJ, with comparable efficiency.

Survey of cyclic voltammograms of lower aliphatic alcohols in 1 M KOH recorded at electrodes containing atomic gold is presented. The atomically structured gold is placed in polyaniline, which is deposited on a platinum electrode. The electrochemical behavior of these atomic gold electrodes (AGE) is compared with Pt/polyaniline electrode without gold and with Pt/polyaniline electrode containing “unstructured” nanoclusters of Au. The conditions were identical for all of the experiments. This allows direct visual comparison of the effect of atomic structuring. The electrochemical activity of the AGEs is dominated by the odd–even pattern of reactivity, and it is superimposed on specific electrochemical behavior of individual alcohols.

The effect of addition of one atom of palladium to n atoms of gold (n = 1–5) dispersed in polyaniline electrodes, for the electrooxidation of methanol and ethanol in alkaline solution has been investigated. For comparison, oxidation at pure atomic metal electrodes, Pt/PANI–Au n=2–7 and Pt/PANI–Pd n=2–6, was performed. It is shown that the one added Pd atom affects the oxidation peak currents and makes a significant difference in selectivity. Similar to our previous studies, the electrocatalytic activity has been correlated with the theoretically predicted HOMO–LUMO gap energies for the atomic metal alloys. Effect of addition of one atom of Pd to Au n in polyaniline .

The previously described cyclic pathway method for deposition of atomic metals has been used to create Pd1–6 atomic clusters and Au1–5Pd1 and Au1–4Pd2 bimetallic atomic clusters in polyaniline (PANI). The controlled deposition of predetermined atomic size clusters of metals has been examined by testing the electrochemical oxidation of n-propanol in 1 M NaOH. The oxidation peak currents from the cyclic voltammograms were found to follow the same trend as the changes of the calculated HOMO–LUMO gap energies. The FTIR signature of PANI for these films also followed the calculated trend. This study also looks at the effects of atomic arrangement in the atomic structure on the support matrix of PANI. The results presented here have shown that the cyclic pathway is a versatile method for the atomic deposition of single metal, bimetallic, or even trimetallic atomic clusters in PANI.

Combinatorial biosynthesis for more efficient antibiotics: 150 novel lincosamides prepared by combining lincomycin and celesticetin biosynthetic pathways., Lincosamides, which are produced by streptomycetes, compose a small but clinically important class of antibiotics. The recent elucidation of the condensation and post-condensation biosynthetic steps of the lincosamides lincomycin and celesticetin revealed several unexpected reaction mechanisms. Here, we prepared recombinant proteins involved in the celesticetin biosynthetic pathway and used them for in vitro assays that were monitored by LC-MS. Our results elucidate the last biosynthetic step of celesticetin: the attachment of salicylic acid is catalyzed by the Ccb2 acyl-CoA ligase and the Ccb1 acyltransferase. Ccb1 belongs to the WS/DGAT protein family and, in contrast to the characterized members of the family, has unusual substrate specificity. To the best of our knowledge, Ccb1 is the first protein in this family that transfers a benzoyl derivative-CoA conjugate and is the first WS/DGAT protein involved in the biosynthesis of secondary metabolites. Furthermore, we exploited the relaxed substrate specificities of Ccb1 and Ccb2, as well as three additional upstream post-condensation biosynthetic proteins in the celesticetin pathway, and combined the lincomycin and the celesticetin biosynthetic pathways in vitro. In this way, we prepared a library of 150 novel hybrid lincosamides, including two unnatural chimeras of lincomycin and celesticetin, which were shown to have antibacterial properties more pronounced than clinically used lincomycin. These achievements may be considered a case study in applying knowledge about biosynthetic machinery to assemble a large number of compounds from originally a small group of natural products without the need for chemical synthesis.

A sorption process of RuCl3 in phosphate buffer by polyaniline (PANI) powder chemically synthesized from phosphoric acid was spectrophotometrically monitored as a function of time. It was determined that the sorption process follows the Langmuir and Freundlich isotherms, and their constants were evaluated. It was determined that chemisorption was the rate-controlling step. By conducting detailed studies, we assigned the chemisorption to Lewis acid based interactions of the sorbent electron pair localized at the benzenoid amine (−NH2) and quinoid imine (═NH) groups, with the sorbate, RuCl3, as the electron acceptor. The stability of the interaction over a period of ∼1 week showed that the presence of the Ru(III) in the PANI matrix reverses its state from emeraldine base to emeraldine salt, resulting in a change of conductivity. The partial electron donor based charge transfer is a slow process as compared to the sorption process involving Bronsted acid doping.

Defined diblock and triblock copolymers composed of methyl methacrylate-co-glycidyl methacrylate block and 3-{3,5,7,9,11,13,15-hepta(2-methylpropyl)-pentacyclo[9.5.1.13,9.15,15.17,13]-octasiloxan-1-yl}propyl methacrylate block(s), i.e., P(MMA-co-GMA)-b-PiBuPOSSMA and PiBuPOSSMA-b-P(MMA-co-GMA)-b-PiBuPOSSMA, were synthesized by atom transfer radical polymerization (ATRP). First, monofunctional and bifunctional P(MMA-co-GMA) copolymers were synthesized by ATRP. Subsequently, these copolymers were successfully used as macroinitiators for ATRP of POSS-containing methacrylate monomer. The process showed high initiation efficiency of macroinitiators and led to products with low dispersity. The synthesized block copolymers were characterized by size exclusion chromatography, 1H-NMR spectroscopy and their glass transition temperatures were determined by differential scanning calorimetry. © 2012 Wiley Periodicals, Inc. J. Appl. Polym. Sci., 2013

Structurally different and functionally diverse natural compounds – antitumour agents pyrrolo[1,4]benzodiazepines, bacterial hormone hormaomycin, and lincosamide antibiotic lincomycin – share a common building unit, 4-alkyl-L-proline derivative (APD). APDs arise from L-tyrosine through a special biosynthetic pathway. Its generally accepted scheme, however, did not comply with current state of knowledge. Based on gene inactivation experiments and in vitro functional tests with recombinant enzymes, we designed a new APD biosynthetic scheme for the model of lincomycin biosynthesis. In the new scheme at least one characteristic in each of five final biosynthetic steps has been changed: the order of reactions, assignment of enzymes and/or reaction mechanisms. First, we demonstrate that LmbW methylates a different substrate than previously assumed. Second, we propose a unique reaction mechanism for the next step, in which a putative γ-glutamyltransferase LmbA indirectly cleaves off the oxalyl residue by transient attachment of glutamate to LmbW product. This unprecedented mechanism would represent the first example of the C–C bond cleavage catalyzed by a γ-glutamyltransferase, i.e., an enzyme that appears unsuitable for such activity. Finally, the inactivation experiments show that LmbX is an isomerase indicating that it transforms its substrate into a compound suitable for reduction by LmbY, thereby facilitating its subsequent complete conversion to APD 4-propyl-L-proline. Elucidation of the APD biosynthesis has long time resisted mainly due to the apparent absence of relevant C–C bond cleaving enzymatic activity. Our proposal aims to unblock this situation not only for lincomycin biosynthesis, but generally for all above mentioned groups of bioactive natural products with biotechnological potential.

Here, we present the first example of application of single-electron transfer living radical polymerization (SET-LRP) process to a controlled grafting of cellulose esters, cellulose diacetate (CDA), and cellulose acetate butyrate (CAB). The cellulose ester macroinitiators with various functionality densities have been prepared by acylation of the backbones with 2-bromoisobutyryl (BrIB) and dichloroacetyl (DCA) groups, respectively. Methacrylate monomers were polymerized using DCA-functionalized macroinitiators in the presence of pentamethyldiethylene triamine as a ligand. At 30 °C, the reaction is rather slow, reaching about 10% conversion after 3 to 6 h of polymerization, whereas the higher temperature (60 °C) perceptibly speeds up the polymerization so that methyl methacrylate (MMA) conversion is ∼30% after 5 h. Graft copolymers with random-type and diblock-type grafts having amphiphilic character were also synthesized. For acrylate grafting (BuA and t-BuA), BrIB-functionalized macroinitiators are more convenient in a combination with a low concentration of Cu(0) and Me6TREN as a ligand and polymerization is detectably faster even at the lower temperature than that of MMA. Kinetic studies show “living” character of both the graftings. Important advantages of SET-LRP, compared with classic ATRP, are (i) higher polymerization rate, (ii) lower extent of recombination of the growing grafts and (iii) negligible coloration of the products with catalytic residua, so that the prepared polymers do not require additional careful purification. © 2010 Wiley Periodicals, Inc. J Polym Sci Part A: Polym Chem, 2010

In numerous epidemiological studies inverse associations between the intake of brassicaceous crops and chronic diseases, such as cancer and cardiovascular diseases, have been demonstrated. Phytochemicals such as carotenoids or phenolic compounds have been indicated to be responsible for this protective health effect. It is known that distinct changes in the content of certain phytochemicals can be triggered by the application of physical elicitors, e.g., UV and gamma irradiation, during production and postharvest operations. In order to determine the effect of short-term UV-B exposure and gamma irradiation as targeted stress treatments on the secondary plant metabolism, sprouts were subjected to three UV-B radiation intensities: 0.075, 0.10 and 0.15 Wh m -2 . Gamma irradiation was performed in dosages of 0.5, 1.0, 3.0, and 6.0 kGy. The results clearly demonstrate that the plant response to short-term and moderate UV-B exposure as well as gamma irradiation is dependent on the physiological plant stage and on the phytochemical compound.

The lincomycin biosynthetic gene lmbX was deleted in Streptomyces lincolnensis ATCC 25466, and deletion of this gene led to abolition of lincomycin production. The results of complementation experiments proved the blockage in the biosynthesis of lincomycin precursor 4-propyl- l -proline. Feeding this mutant strain with precursor derivatives resulted in production of 4′-butyl-4′-depropyllincomycin and 4′-pentyl-4′-depropyllincomycin in high titers and without lincomycin contamination. Moreover, 4′-pentyl-4′-depropyllincomycin was found to be more active than lincomycin against clinical Staphylococcus isolates with genes determining low-level lincosamide resistance.

This article describes the synthesis and characterization of new amphiphilic polymer conetworks containing hydrophilic poly(methacrylic acid) (PMAA) or poly(acrylic acid) (PAA) and hydrophobic polyisobutylene (PIB) chains. These conetworks were prepared by a two-step polymer synthesis. In the first step, a cationic copolymer of isobutylene (IB) and 3-isopropenyl-α,α-dimethylbenzyl isocyanate (IDI) was prepared. The isocyanate groups of the IB–IDI random copolymer were subsequently transformed in situ to methacrylate (MA) groups in reaction with 2-hydroxyethyl methacrylate (HEMA). In the second step, the resulting MA-multifunctional PIB-based crosslinker, PIB(MA)n, with an average functionality of approximately four methacrylic groups per chain, was copolymerized with methacrylic acid (MAA) or acrylic acid (AA) by radical mechanism in tetrahydrofuran giving rise to amphiphilic conetworks containing 31–79 mol % of MAA or 26–36 mol % of AA. The synthesized conetworks were characterized with solid-state 13C-NMR spectroscopy and differential scanning calorimetry. The amphiphilic nature of the conetworks was proven by swelling in both aqueous media with low and high pH and n-heptane. The effect of varying pH on the swelling behavior of the synthesized conetworks is presented. © 2009 Wiley Periodicals, Inc. J Polym Sci Part A: Polym Chem 47: 1284–1291, 2009

Field-effect transistors with mixed ionic-electronic conductors have been prepared by varying the ratio of room temperature ionic liquid and emeraldine salt of polyaniline. Transistor with sufficiently high electronic conductivity (32 mol% ES-PANI) and Au gate contact exhibited theoretical behavior of insulated gate field-effect transistor. On the other hand the purely ionic gate behaved irreproducibly, indicating that a capacitive divider has been formed in the gate.

Experimental realization of a reference potential for potentiometric measurements in non-ideal media is discussed. A liquid junction-free reference potential is realized at a dropping mercury electrode, using the standard potential of the metallocene redox couple. This approach is particularly suitable for determination of acidity functions that are seamlessly anchored to aqueous standard buffers.

Acetohydroxy-acid synthases (AHAS) of two mutant strains Streptomyces cinnamonensis ACB-NLR-2 and BVR-18 were chosen for this study for their apparent activation by valine, which regularly acts as an allosteric inhibitor. Sequencing the ilvB genes coding for the AHAS catalytic subunit revealed two distant changes in the mutants, DeltaQ217 and E139A, respectively. Homology modeling was used to propose the structural changes caused by those mutations. In the mutant strain ACB-NLR-2 (resistant to 2-amino-3-chlorobutyrate and norleucine), deletion of Q217 affected a helix in ss-domain, distant from the active center. As no mutation was found in the regulatory subunit of this strain, DeltaQ217 in IlvB was supposed to be responsible for the observed valine activation, probably via changed properties on the proposed regulatory-catalytic subunit interface. In mutant strain BVR-18 (resistant to 2-oxobutyrate), substitution E139A occurred in a conservative loop near the active center. In vitro AHAS activity assay with the enzyme reconstituted from the wild-type regulatory and BVR-18 catalytic subunits proved that the substitution in the catalytic subunit led to the apparent activation of AHAS by valine. We suggest that the conservative loop participated in a conformational change transfer to the active center during the allosteric regulation.

Polyaniline forms a strong complex with chloroaurate at the protonated imine sites. Here we report on an electrochemical procedure that allows preparation of gold clusters by adding gold atoms one-by-one (“bottom–up” approach). It is contrasted with the “top–down” approach in which the growth of multi-atom Au clusters was also controlled electrochemically. Our results confirm that both the amount and the size of gold clusters affect the properties of the composite material.

The product of spontaneous termination formed after addition of one or two t-BuA units onto living PMMA chains in the MMA/t-BuA block copolymerization was isolated and characterized by SEC, UV, FT-IR, Raman and NMR spectroscopy. It appears as a low-molecular-weight peak in SEC eluograms of the copolymers, absorbing at 260 nm; its retention time corresponds to that of the PMMA block. In its FT-IR and Raman spectra, new bands appeared corresponding to the C=C and C=O vibrations of a conjugated and H-bonded ester group of the enol form of the cyclic oxoester composed of MMA and t-BuA units. Experimental support of a back-biting reaction at the link between PMMA and Pt-BuA blocks is presented.

The rate constants of the reactions of eaq− and the OH· radical with the oxalate ion in a neutral aqueous solution were measured by means of the pulse radiolysis technique. They were found to be (3.5 ± 0.5) × 107 and (1.5 ± 0.2) × 107 l mol−1 s−1, respectively. The radical anion −OOC-C·OO2− is characterized by an optical absorption band that has a maximum at 270 nm and a molar absorption coefficient of (2400 ± 200) l mol−1 cm−1. The radical anion ·OOC-COO−, the product of the reaction with the OH· radical, exhibits absorption that has no maximum and increases in intensity with a decrease in the wavelength extending to the UV region (ɛ220 = 1800 l mol−1 cm−1). The mechanism of radiation-chemical transformations in aqueous oxalate solutions is discussed.

We determined the activities of new antibiotics telithromycin (ketolide) and quinupristin/dalfo- pristin (streptogramins) against 88 macrolide and/or lincosamide resistant coagulase-negative staphylococci (CoNS) isolates with defined resistance gene status. Telithromycin susceptibility was determined only in erythromycin-sensitive isolates (15) indicating the same mechanisms of resistance. In contrast, all erythro- mycin-resistant isolates (73) were either constitutively resistant to telithromycin (13 isolates with constitu- tive erm genes) or demonstrated telithromycin D-shaped zone (60 isolates with inducible msr(A) and/or erm). However, the level of inducible resistance conferred by msr(A) (35 isolates) was borderline even after induction by erythromycin. No quinupristin/dalfopristin resistant isolate was observed if tested by disk-dif- fusion method (DDM) but 18 isolates were intermediate (MIC = 1-3 mg/L) and two isolates resistant (MIC = 8 mg/L) if tested by E-test. All these isolates were resistant to streptogramin A and harbored vga(A) gene (1 isolate) or vga(A)LC gene (19 isolates). MICs for quinupristin/dalfopristin were higher for isolates with combination of streptogramin A resistance and constitutive MLSB resistance (MIC = 3-8 mg/L in 4 isolates) than for streptogramin A-resistant isolates susceptible to streptogramin B (MIC = 0.5-2 mg/L in 16 isolates). In addition to S. haemolyticus, vga(A)LC was newly identified in S. epidermidis and S. warnerii indicating its widespread occurrence in CoNS. Misidentification of low-level resistant isolates by DDM may contribute to dissemination of streptogramin A resistance.

Bifunctional polystyrene macroinitiators, having various molecular weights, were prepared by atom transfer radical polymerization (ATRP), initiated with bifunctional initiator 1,3-bis{1-methyl-1[(2,2,2-trichloroethoxy) carbonylamino]ethyl}benzene in conjunction with CuCl catalyst and polyamine ligands. These macroinitiators were subsequently used for ATRP of tert-butyl acrylate (t-BuA), giving BAB triblocks poly[(t-BuA)-b-(Sty)-b-(t-BuA)] as precursors of amphiphilic copolymers. Both the polymerization steps proceeded as controlled processes with linear semi-logarithmic conversion plots and lengths of the blocks following theoretical predictions. Hydrolysis of outer poly(t-BuA) blocks led to triblock copolymers with the central polystyrene block and outer blocks of poly(acrylic acid), the molecular weights of which ranged from ca. 5 × 103 to almost 1 × 105 Da.

The first insight into celesticetin biosynthetic gene cluster of S. caelestis is presented. The genomic DNA of producing strain was digested, digoxigenin-labeled and hybridized with a set of probes designed according to S. lincolnensis gene sequences. Genes with high homology to the lincomycin biosynthetic genes coding for the predicted common parts of the pathway were identified in S. caelestis. Then, genomic DNA of S. caelestis treated by a multiple digestion was hybridized with five digoxigenin-labeled probes to construct a rough restriction map. Two consecutive islands formed by the genes with a putative function in biosynthesis of the shared saccharide moiety revealed an organization similar to the lincomycin biosynthetic gene cluster. The celesticetin cluster was mapped and essential information was obtained for subsequent steps, i.e. isolation and sequence analysis of the cluster.

Three examples from the field of physical chemistry are given here in order to demonstrate the versatility of optical pulse radiolysis measurements in the UV region: (i) the formation and disappearance of O − in oxygen-free solution at pH 13 has been studied again and its spectrum has been re-determined, (ii) because the value of the rate constant for the reaction of hydrogen atoms with tert-butyl alcohol became ambiguous during the last years, direct optical measurement in the UV region were carried out in which the build-up of the absorption of the subsequently formed alcohol radical was followed directly, (iii) in some preliminary experiments, fused silica (Suprasil II) was irradiated by trains of high-energy electron pulses and the subsequent absorption was recorded.