Open AccessArticle
Extended Physicochemical Characterization of the Synthetic Anticoagulant Pentasaccharide Fondaparinux Sodium by Quantitative NMR and Single Crystal X-ray Analysis
Molecules 2017, 22(8), 1362; doi:10.3390/molecules22081362 (registering DOI) -
Abstract
Fondaparinux sodium is a synthetic pentasaccharide representing the high affinity antithrombin III binding site in heparin. It is the active pharmaceutical ingredient of the anticoagulant drug Arixtra®. The single crystal X-ray structure of Fondaparinux sodium is reported, unequivocally confirming both structure
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Fondaparinux sodium is a synthetic pentasaccharide representing the high affinity antithrombin III binding site in heparin. It is the active pharmaceutical ingredient of the anticoagulant drug Arixtra®. The single crystal X-ray structure of Fondaparinux sodium is reported, unequivocally confirming both structure and absolute configuration. The iduronic acid adopts a somewhat distorted chair conformation. Due to the presence of many sulfur atoms in the highly sulfated pentasaccharide, anomalous dispersion could be applied to determine the absolute configuration. A comparison with the conformation of Fondaparinux in solution, as well as complexed with proteins is presented. The content of the solution reference standard was determined by quantitative NMR using an internal standard both in 1999 and in 2016. A comparison of the results allows the conclusion that this method shows remarkable precision over time, instrumentation and analysts. Full article
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Open AccessArticle
Spirulina maxima Extract Prevents Neurotoxicity via Promoting Activation of BDNF/CREB Signaling Pathways in Neuronal Cells and Mice
Molecules 2017, 22(8), 1363; doi:10.3390/molecules22081363 (registering DOI) -
Abstract
Spirulina maxima is a microalgae which contains flavonoids and other polyphenols. Although Spirulina maxima 70% ethanol extract (SM70EE) has diverse beneficial effects, its effects on neurotoxicity have not been fully understood. In this study, we investigated the neuroprotective effects of SM70EE against trimethyltin
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Spirulina maxima is a microalgae which contains flavonoids and other polyphenols. Although Spirulina maxima 70% ethanol extract (SM70EE) has diverse beneficial effects, its effects on neurotoxicity have not been fully understood. In this study, we investigated the neuroprotective effects of SM70EE against trimethyltin (TMT)-induced neurotoxicity in HT-22 cells. SM70EE inhibited the cleavage of poly-ADP ribose polymerase (PARP). Besides, ROS production was decreased by down-regulating oxidative stress-associated enzymes. SM70EE increased the factors of brain-derived neurotrophic factor (BDNF)/cyclic AMPresponsive elementbinding protein (CREB) signalling pathways. Additionally, acetylcholinesterase (AChE) was suppressed by SM70EE. Furthermore, we investigated whether SM70EE prevents cognitive deficits against scopolamine-induced neurotoxicity in mice by applying behavioral tests. SM70EE increased step-through latency time and decreased the escape latency time. Therefore, our data suggest that SM70EE may prevent TMT neurotoxicity through promoting activation of BDNF/CREB neuroprotective signaling pathways in neuronal cells. In vivo study, SM70EE would prevent cognitive deficits against scopolamine-induced neurotoxicity in mice. Full article
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Open AccessArticle
Biochemical and Comparative Transcriptomic Analyses Identify Candidate Genes Related to Variegation Formation in Paeonia rockii
Molecules 2017, 22(8), 1364; doi:10.3390/molecules22081364 (registering DOI) -
Abstract
Paeonia rockii is a wild tree peony species with large and dark purple variegations at the base of its petals. It is the genetic resource for various variegation patterns in tree peony cultivars, which is in contrast to the pure white petals of
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Paeonia rockii is a wild tree peony species with large and dark purple variegations at the base of its petals. It is the genetic resource for various variegation patterns in tree peony cultivars, which is in contrast to the pure white petals of Paeonia ostii. However, the molecular mechanism underlying the formation of variegation in this plant is still unknown. Here, we conducted Illumina transcriptome sequencing for P. rockii, P. ostii (with pure white petals) and their F1 individuals (with purple-red variegation). A total of 181,866 unigenes were generated, including a variety of unigenes involved in anthocyanin biosynthesis and sequestration and the regulation of anthocyanin biosynthesis. The dark purple or purple-red variegation patterns mainly occurred due to the proportions of cyanidin (Cy)- and peonidin (Pn)-based anthocyanins. The variegations of P. rockii exhibited a “Cy > Pn” phenotype, whereas the F1 progeny showed a “Pn > Cy” phenotype. The CHS, DFR, ANS, and GST genes might play key roles in variegation pigmentation in P. rockii according to gene expression and interaction network analysis. Two R2R3-MYB transcription factors (c131300.graph_c0 and c133735.graph_c0) regulated variegation formation by controlling CHS, ANS and GST genes. Our results indicated that the various variegation patterns were caused by transcriptional regulation of anthocyanin biosynthesis genes, and the transcription profiles of the R2R3-MYBs provided clues to elucidate the mechanisms underlying this trait. The petal transcriptome data produced in this study will provide a valuable resource for future association investigations of the genetic regulation of various variegation patterns in tree peonies. Full article
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Open AccessArticle
Anticancer Activity of Ramalin, a Secondary Metabolite from the Antarctic Lichen Ramalina terebrata, against Colorectal Cancer Cells
Molecules 2017, 22(8), 1361; doi:10.3390/molecules22081361 (registering DOI) -
Abstract
Colorectal cancer is a leading cause of death worldwide and occurs through the highly complex coordination of multiple cellular pathways, resulting in carcinogenesis. Recent studies have increasingly revealed that constituents of lichen extracts exhibit potent pharmaceutical activities, including anticancer activity against various cancer
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Colorectal cancer is a leading cause of death worldwide and occurs through the highly complex coordination of multiple cellular pathways, resulting in carcinogenesis. Recent studies have increasingly revealed that constituents of lichen extracts exhibit potent pharmaceutical activities, including anticancer activity against various cancer cells, making them promising candidates for new anticancer therapeutic drugs. The main objective of this study was to evaluate the anticancer capacities of ramalin, a secondary metabolite from the Antarctic lichen Ramalina terebrata, in the human colorectal cancer cell line HCT116. In this study, ramalin displayed concentration-dependent anticancer activity against HCT116 cells, significantly suppressing proliferation and inducing apoptosis. Furthermore, ramalin induced cell cycle arrest in the gap 2/mitosis (G2/M) phase through the modulation of hallmark genes involved in the G2/M phase transition, such as tumour protein p53 (TP53), cyclin-dependent kinase inhibitor 1A (CDKN1A), cyclin-dependent kinase 1 (CDK1) and cyclin B1 (CCNB1). At both the transcriptional and translational level, ramalin caused a gradual increase in the expression of TP53 and its downstream gene CDKN1A, while decreasing the expression of CDK1 and CCNB1 in a concentration-dependent manner. In addition, ramalin significantly inhibited the migration and invasion of colorectal cancer cells in a concentration-dependent manner. Taken together, these data suggest that ramalin may be a therapeutic candidate for the targeted therapy of colorectal cancer. Full article
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Open AccessReview
Bioactive Compounds from Mexican Varieties of the Common Bean (Phaseolus vulgaris): Implications for Health
Molecules 2017, 22(8), 1360; doi:10.3390/molecules22081360 (registering DOI) -
Abstract
As Mexico is located within Mesoamerica, it is considered the site where the bean plant originated and where it was domesticated. Beans have been an integral part of the Mexican diet for thousands of years. Within the country, there are a number of
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As Mexico is located within Mesoamerica, it is considered the site where the bean plant originated and where it was domesticated. Beans have been an integral part of the Mexican diet for thousands of years. Within the country, there are a number of genotypes possessing highly diverse physical and chemical properties. This review describes the major bioactive compounds contained on the Mexican varieties of the common bean. A brief analysis is carried out regarding the benefits they have on health. The effect of seed coat color on the nutraceutical compounds content is distinguished, where black bean stands out because it is high content of anthocyanins, polyphenols and flavonoids such as quercetin. This confers black bean with an elevated antioxidant capacity. The most prominent genotypes within this group are the “Negro San Luis”, “Negro 8025” and “Negro Jamapa” varieties. Conversely, the analyzed evidence shows that more studies are needed in order to expand our knowledge on the nutraceutical quality of the Mexican bean genotypes, either grown or wild-type, as well as their impact on health in order to be used in genetic improvement programs or as a strategy to encourage their consumption. The latter is based on the high potential it has for health preservation and disease prevention. Full article
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Open AccessFeature PaperArticle
Lactose Binding Induces Opposing Dynamics Changes in Human Galectins Revealed by NMR-Based Hydrogen–Deuterium Exchange
Molecules 2017, 22(8), 1357; doi:10.3390/molecules22081357 -
Abstract
Galectins are β-galactoside-binding proteins implicated in a myriad of biological functions. Despite their highly conserved carbohydrate binding motifs with essentially identical structures, their affinities for lactose, a common galectin inhibitor, vary significantly. Here, we aimed to examine the molecular basis of differential lactose
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Galectins are β-galactoside-binding proteins implicated in a myriad of biological functions. Despite their highly conserved carbohydrate binding motifs with essentially identical structures, their affinities for lactose, a common galectin inhibitor, vary significantly. Here, we aimed to examine the molecular basis of differential lactose affinities amongst galectins using solution-based techniques. Consistent dissociation constants of lactose binding were derived from nuclear magnetic resonance (NMR) spectroscopy, intrinsic tryptophan fluorescence, isothermal titration calorimetry and bio-layer interferometry for human galectin-1 (hGal1), galectin-7 (hGal7), and the N-terminal and C-terminal domains of galectin-8 (hGal8NTD and hGal8CTD, respectively). Furthermore, the dissociation rates of lactose binding were extracted from NMR lineshape analyses. Structural mapping of chemical shift perturbations revealed long-range perturbations upon lactose binding for hGal1 and hGal8NTD. We further demonstrated using the NMR-based hydrogen–deuterium exchange (HDX) that lactose binding increases the exchange rates of residues located on the opposite side of the ligand-binding pocket for hGal1 and hGal8NTD, indicative of allostery. Additionally, lactose binding induces significant stabilisation of hGal8CTD across the entire domain. Our results suggested that lactose binding reduced the internal dynamics of hGal8CTD on a very slow timescale (minutes and slower) at the expense of reduced binding affinity due to the unfavourable loss of conformational entropy. Full article
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Open AccessArticle
Insights into the Effect of the G245S Single Point Mutation on the Structure of p53 and the Binding of the Protein to DNA
Molecules 2017, 22(8), 1358; doi:10.3390/molecules22081358 -
Abstract
The transcription factor p53 is a potent tumor suppressor dubbed as the “guardian of the genome” because of its ability to orchestrate protective biological outputs in response to a variety of oncogenic stresses. Mutation and thus inactivation of p53 can be found in
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The transcription factor p53 is a potent tumor suppressor dubbed as the “guardian of the genome” because of its ability to orchestrate protective biological outputs in response to a variety of oncogenic stresses. Mutation and thus inactivation of p53 can be found in 50% of human tumors. The majority are missense mutations located in the DNA binding region. Among them, G245S is known to be a structural hotspot mutation. To understand the behaviors and differences between the wild-type and mutant, both a dimer of the wild type p53 (wt-p53) and its G245S mutant (G245S-mp53), complexed with DNA, were simulated using molecular dynamics for more than 1 μs. wt-p53 and G245S-mp53 apo monomers were simulated for 1 μs as well. Conformational analyses and binding energy evaluations performed underline important differences and therefore provide insights to understand the G245S-mp53 loss of function. Our results indicate that the G245S mutation destabilizes several structural regions in the protein that are crucial for DNA binding when found in its apo form and highlight differences in the mutant-DNA complex structure compared to the wt protein. These findings not only provide means that can be applied to other p53 mutants but also serve as structural basis for further studies aimed at the development of cancer therapies based on restoring the function of p53. Full article
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Open AccessArticle
Transcriptional Responses of Creeping Bentgrass to 2,3-Butanediol, a Bacterial Volatile Compound (BVC) Analogue
Molecules 2017, 22(8), 1318; doi:10.3390/molecules22081318 -
Abstract
Bacterial volatile compounds (BVCs) have been reported to enhance plant growth and elicit plant defenses against fungal infection and insect damage. The objective of this study was to determine transcriptomic changes in response to synthetic BVC that could be associated with plant resistance
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Bacterial volatile compounds (BVCs) have been reported to enhance plant growth and elicit plant defenses against fungal infection and insect damage. The objective of this study was to determine transcriptomic changes in response to synthetic BVC that could be associated with plant resistance to Rhizoctonia solani in creeping bentgrass. The 2,3-butanediol (BD) (250 µM) was sprayed on creeping bentgrass leaves grown in jam jars. The result showed that synthetic BD induced plant defense against R. solani for creeping bentgrass. Transcriptomic analysis demonstrated that more genes were repressed by BD while less showed up-regulation. BD suppressed the expression of some regular stress-related genes in creeping bentgrass, such as pheromone activity, calcium channel activity, photosystem II oxygen evolving complex, and hydrolase activity, while up-regulated defense related transcription factors (TFs), such as basic helix-loop-helix (bHLH) TFs, cysteine2-cysteine2-contans-like (C2C2-CO) and no apical meristem TFs (NAC). Other genes related to disease resistance, such as jasmonic acid (JA) signaling, leucine rich repeats (LRR)-transmembrane protein kinase, pathogen-related (PR) gene 5 receptor kinase and nucleotide binding site-leucine rich repeats (NBS-LRR) domain containing plant resistance gene (R-gene) were also significantly up-regulated. These results suggest that BD may induce changes to the plant transcriptome in induced systemic resistance (ISR) pathways. Full article
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Open AccessArticle
Sulfonamide-Linked Ciprofloxacin, Sulfadiazine and Amantadine Derivatives as a Novel Class of Inhibitors of Jack Bean Urease; Synthesis, Kinetic Mechanism and Molecular Docking
Molecules 2017, 22(8), 1352; doi:10.3390/molecules22081352 -
Abstract
Sulfonamide derivatives serve as an important building blocks in the drug design discovery and development (4D) process. Ciprofloxacin-, sulfadiazine- and amantadine-based sulfonamides were synthesized as potent inhibitors of jack bean urease and free radical scavengers. Molecular diversity was explored and electronic factors were
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Sulfonamide derivatives serve as an important building blocks in the drug design discovery and development (4D) process. Ciprofloxacin-, sulfadiazine- and amantadine-based sulfonamides were synthesized as potent inhibitors of jack bean urease and free radical scavengers. Molecular diversity was explored and electronic factors were also examined. All 24 synthesized compounds exhibited excellent potential against urease enzyme. Compound 3e (IC50 = 0.081 ± 0.003 µM), 6a (IC50 = 0.0022 ± 0.0002 µM), 9e (IC50 = 0.0250 ± 0.0007 µM) and 12d (IC50 = 0.0266 ± 0.0021 µM) were found to be the lead compounds compared to standard (thiourea, IC50 = 17.814 ± 0.096 µM). Molecular docking studies were performed to delineate the binding affinity of the molecules and a kinetic mechanism of enzyme inhibition was propounded. Compounds 3e, 6a and 12d exhibited a mixed type of inhibition, while derivative 9e revealed a non-competitive mode of inhibition. Compounds 12a, 12b, 12d, 12e and 12f showed excellent radical scavenging potency in comparison to the reference drug vitamin C. Full article
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Open AccessReview
Application of Solution NMR to Structural Studies on α-Helical Integral Membrane Proteins
Molecules 2017, 22(8), 1347; doi:10.3390/molecules22081347 -
Abstract
A large portion of proteins in living organisms are membrane proteins which play critical roles in the biology of the cell, from maintenance of the biological membrane integrity to communication of cells with their surroundings. To understand their mechanism of action, structural information
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A large portion of proteins in living organisms are membrane proteins which play critical roles in the biology of the cell, from maintenance of the biological membrane integrity to communication of cells with their surroundings. To understand their mechanism of action, structural information is essential. Nevertheless, structure determination of transmembrane proteins is still a challenging area, even though recently the number of deposited structures of membrane proteins in the PDB has rapidly increased thanks to the efforts using X-ray crystallography, electron microscopy, and solid and solution nuclear magnetic resonance (NMR) technology. Among these technologies, solution NMR is a powerful tool for studying protein-protein, protein-ligand interactions and protein dynamics at a wide range of time scales as well as structure determination of membrane proteins. This review provides general and useful guideline for membrane protein sample preparation and the choice of membrane-mimetic media, which are the key step for successful structural analysis. Furthermore, this review provides an opportunity to look at recent applications of solution NMR to structural studies on α-helical membrane proteins through some success stories. Full article
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Open AccessCommunication
Fenton Discoloration of Ultrasonicated Purple Cactus Pear Juice
Molecules 2017, 22(8), 1344; doi:10.3390/molecules22081344 -
Abstract
The aim of this study was to evaluate the stability of color, betaxanthin, and betacyanin pigments in the presence of Cu(II)-dependent hydroxyl radicals (HO•) from ultrasonicated purple cactus pear juice at amplitudes of 40%, 60%, and 80%, in comparison to untreated sample. L*
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The aim of this study was to evaluate the stability of color, betaxanthin, and betacyanin pigments in the presence of Cu(II)-dependent hydroxyl radicals (HO•) from ultrasonicated purple cactus pear juice at amplitudes of 40%, 60%, and 80%, in comparison to untreated sample. L* parameter of juice treated at 40% and 80% amplitude for 25 and 15 min, respectively (11.3 and 9.3, respectively), were significantly higher compared to the control; b* and hue parameters of juice treated at 80%, 25 min showed values of 1.7 and 0.1, respectively. Color differences (ΔE) were lower (<3) for juices treated at high amplitude (80%) and short times (3–5 min). Juice treated at 40% 15 min, 60% 25 min, 80% 15 and 25 min presented high values of betacyanins (281.7 mg·L−1, 255.9 mg·L−1, 294.4 mg·L−1, and 276.7 mg·L−1, respectively). Betaxanthin values were higher in the juices treated at 40% 5 min and 80% 15 and 25 min (154.2 mg·L−1, 135.2 mg·L−1, and 128.5 mg·L−1, respectively). Purple cactus pear juice exhibited significant chelating activity of copper ions and great stability when exposed to HO•. Full article
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Open AccessArticle
Effect of 2,6-Bis-(1-hydroxy-1,1-diphenyl-methyl) Pyridine as Organic Additive in Sulfide NiMoP/γ-Al2O3 Catalyst for Hydrodesulfurization of Straight-Run Gas Oil
Molecules 2017, 22(8), 1332; doi:10.3390/molecules22081332 -
Abstract
The effect of 2,6-bis-(1-hydroxy-1,1-diphenyl-methyl) pyridine (BDPHP) in the preparation of NiMoP/γ-Al2O3 catalysts have been investigated in the hydrodesulfurization (HDS) of straight-run gas oil. The γ-Al2O3 support was modified by surface impregnation of a solution of BDPHP to
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The effect of 2,6-bis-(1-hydroxy-1,1-diphenyl-methyl) pyridine (BDPHP) in the preparation of NiMoP/γ-Al2O3 catalysts have been investigated in the hydrodesulfurization (HDS) of straight-run gas oil. The γ-Al2O3 support was modified by surface impregnation of a solution of BDPHP to afford BDPHP/Ni molar ratios (0.5 and 1.0) in the final composition. The highest activity for NiMoP materials was found when the molar ratio of BDPHP/Ni was of 0.5. X-ray diffraction (XRD) results revealed that NiMoP (0.5) showed better dispersion of MoO3 than the NiMoP (1.0). Fourier transform infrared spectroscopy (FT-IR) results indicated that the organic additive interacts with the γ-Al2O3 surface and therefore discards the presence of Mo or Ni complexes. Raman spectroscopy suggested a high Raman ratio for the NiMoP (0.5) sample. The increment of the Mo=O species is related to a major availability of Mo species in the formation of MoS2. The temperature programmed reduction (TPR) results showed that the NiMoP (0.5) displayed moderate metal–support interaction. Likewise, X-ray photoelectron spectroscopy (XPS) exhibited higher sulfurization degree for NiMoP (0.5) compared with NiMoP (1.0). The increment of the MoO3 dispersion, the moderate metal–support interaction, the increase of sulfurization degree and the increment of Mo=O species provoked by the BDPHP incorporation resulted in a higher gas oil HDS activity. Full article
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Open AccessArticle
A Unique and Simple Approach to Improve Sensitivity in 15N-NMR Relaxation Measurements for NH3+ Groups: Application to a Protein-DNA Complex
Molecules 2017, 22(8), 1355; doi:10.3390/molecules22081355 -
Abstract
NMR spectroscopy is a powerful tool for research on protein dynamics. In the past decade, there has been significant progress in the development of NMR methods for studying charged side chains. In particular, NMR methods for lysine side-chain NH3+ groups have
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NMR spectroscopy is a powerful tool for research on protein dynamics. In the past decade, there has been significant progress in the development of NMR methods for studying charged side chains. In particular, NMR methods for lysine side-chain NH3+ groups have been proven to be powerful for investigating the dynamics of hydrogen bonds or ion pairs that play important roles in biological processes. However, relatively low sensitivity has been a major practical issue in NMR experiments on NH3+ groups. In this paper, we present a unique and simple approach to improve sensitivity in 15N relaxation measurements for NH3+ groups. In this approach, the efficiency of coherence transfers for the desired components are maximized, whereas undesired anti-phase or multi-spin order components are purged through pulse schemes and rapid relaxation. For lysine side-chain NH3+ groups of a protein-DNA complex, we compared the data obtained with the previous and new pulse sequences under the same conditions and confirmed that the 15N relaxation parameters were consistent for these datasets. While retaining accuracy in measuring 15N relaxation, our new pulse sequences for NH3+ groups allowed an 82% increase in detection sensitivity of 15N longitudinal and transverse relaxation measurements. Full article
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Open AccessArticle
Molecular Mechanisms Underlying Inhibitory Binding of Alkylimidazolium Ionic Liquids to Laccase
Molecules 2017, 22(8), 1353; doi:10.3390/molecules22081353 -
Abstract
Water-miscible alkylimidazolium ionic liquids (ILs) are “green” co-solvents for laccase catalysis, but generally inhibit enzyme activity. Here, we present novel insights into inhibition mechanisms by a combination of enzyme kinetics analysis and molecular simulation. Alkylimidazolium cations competitively bound to the TI Cu
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Water-miscible alkylimidazolium ionic liquids (ILs) are “green” co-solvents for laccase catalysis, but generally inhibit enzyme activity. Here, we present novel insights into inhibition mechanisms by a combination of enzyme kinetics analysis and molecular simulation. Alkylimidazolium cations competitively bound to the TI Cu active pocket in the laccase through hydrophobic interactions. Cations with shorter alkyl chains (C2~C6) entered the channel inside the pocket, exhibiting a high compatibility with laccase (competitive inhibition constant Kic = 3.36~3.83 mM). Under the same conditions, [Omim]Cl (Kic = 2.15 mM) and [Dmim]Cl (Kic = 0.18 mM) with longer alkyl chains bound with Leu296 or Leu297 near the pocket edge and Leu429 around TI Cu, which resulted in stronger inhibition. Complexation with alkylimidazolium cations shifted the pH optima of laccase to the right by 0.5 unit, and might, thereby, lead to invalidation of the Hofmeister series of anions. EtSO4 showed higher biocompatibility than did Ac or Cl, probably due to its binding near the TI Cu and its hindering the entry of alkylimidazolium cations. In addition, all tested ILs accelerated the scavenging of 2, 2′-azino-bis-(3-ethylbenzothiazoline-6-sulphonic acid) (ABTS) radicals, which, however, did not play a determining role in the inhibition of laccase. Full article
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Open AccessArticle
Synthesis and Physicochemical Characterization of the Process-Related Impurities of Eplerenone, an Antihypertensive Drug
Molecules 2017, 22(8), 1354; doi:10.3390/molecules22081354 -
Abstract
Two unknown impurities were observed during the process development for multigram-scale synthesis of eplerenone (Inspra®). The new process-related impurities were identified and fully characterized as the corresponding (7β,11α,17α)-11-hydroxy- and (7α,11β,17α)-9,11-dichloroeplerenone derivatives 12a and 13. Seven other known but poorly described
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Two unknown impurities were observed during the process development for multigram-scale synthesis of eplerenone (Inspra®). The new process-related impurities were identified and fully characterized as the corresponding (7β,11α,17α)-11-hydroxy- and (7α,11β,17α)-9,11-dichloroeplerenone derivatives 12a and 13. Seven other known but poorly described in the literature eplerenone impurities, including four impurities A, B, C and E listed in the European Pharmacopoeia 8.4 were also detected, identified and fully characterized. All these contaminants result from side reactions taking place on the steroid ring C of the starting 11α-hydroxy-7α-(methoxycarbonyl)-3-oxo-17α-pregn-4-ene-21,17-carbolactone (12) and the key intermediate (7α,17α)-9(11)-enester 7, including epimerization of the C-7 asymmetric center, oxidation, dehydration, chlorination and lactonization. The impurities were isolated and/or synthesized and fully characterized by infrared spectroscopy (IR), nuclear magnetic resonance spectroscopy (NMR) and high-resolution mass spectrometry/electrospray ionization (HRMS/ESI). Their 1H- and 13C-NMR signals were fully assigned. The molecular structures of the eight impurities, including the new (7β,11α,17α)-11-hydroxy- and (7α,11β,17α)-9,11-dichloroeplerenone related substances 12a and 13, were solved and refined using single-crystal X-ray diffraction (SCXRD). The full identification and characterization of these impurities should be useful for the quality control and the validation of the analytical methods in the manufacture of eplerenone. Full article
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Open AccessCommunication
Synthesis of 5′-GalNAc-Conjugated Oligonucleotides: A Comparison of Solid and Solution-Phase Conjugation Strategies
Molecules 2017, 22(8), 1356; doi:10.3390/molecules22081356 -
Abstract
Antisense oligonucleotides (ASOs) conjugated to triantennary N-acetyl galactosamine (GalNAc) ligands represent an emerging approach to antisense therapy. Our current generation of GalNAc-ASO conjugates link the GalNAc to the 5′-terminus of the ASO. The conjugation reaction can be accomplished using solution-phase or solid-phase
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Antisense oligonucleotides (ASOs) conjugated to triantennary N-acetyl galactosamine (GalNAc) ligands represent an emerging approach to antisense therapy. Our current generation of GalNAc-ASO conjugates link the GalNAc to the 5′-terminus of the ASO. The conjugation reaction can be accomplished using solution-phase or solid-phase techniques. Here we show a direct comparison of a solution-phase and a solid-phase conjugation strategy. The solution-phase approach, using amine-pentafluorophenyl (PFP) ester coupling, is higher yielding and gives material of slightly higher purity, but requires several additional unit operations and longer production time. The solid-phase approach, using a protected GalNAc ligand phosphoramidite, is more expedient, but results in lower yield and purity. Both strategies efficiently deliver conjugated material in excellent purity. Full article
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Open AccessArticle
Systems Biology Approach to Bioremediation of Nitroaromatics: Constraint-Based Analysis of 2,4,6-Trinitrotoluene Biotransformation by Escherichia coli
Molecules 2017, 22(8), 1242; doi:10.3390/molecules22081242 -
Abstract
Microbial remediation of nitroaromatic compounds (NACs) is a promising environmentally friendly and cost-effective approach to the removal of these life-threating agents. Escherichia coli (E. coli) has shown remarkable capability for the biotransformation of 2,4,6-trinitro-toluene (TNT). Efforts to develop E. coli as
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Microbial remediation of nitroaromatic compounds (NACs) is a promising environmentally friendly and cost-effective approach to the removal of these life-threating agents. Escherichia coli (E. coli) has shown remarkable capability for the biotransformation of 2,4,6-trinitro-toluene (TNT). Efforts to develop E. coli as an efficient TNT degrading biocatalyst will benefit from holistic flux-level description of interactions between multiple TNT transforming pathways operating in the strain. To gain such an insight, we extended the genome-scale constraint-based model of E. coli to account for a curated version of major TNT transformation pathways known or evidently hypothesized to be active in E. coli in present of TNT. Using constraint-based analysis (CBA) methods, we then performed several series of in silico experiments to elucidate the contribution of these pathways individually or in combination to the E. coli TNT transformation capacity. Results of our analyses were validated by replicating several experimentally observed TNT degradation phenotypes in E. coli cultures. We further used the extended model to explore the influence of process parameters, including aeration regime, TNT concentration, cell density, and carbon source on TNT degradation efficiency. We also conducted an in silico metabolic engineering study to design a series of E. coli mutants capable of degrading TNT at higher yield compared with the wild-type strain. Our study, therefore, extends the application of CBA to bioremediation of nitroaromatics and demonstrates the usefulness of this approach to inform bioremediation research. Full article
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Open AccessArticle
ADP-ribosyl-N3: A Versatile Precursor for Divergent Syntheses of ADP-ribosylated Compounds
Molecules 2017, 22(8), 1346; doi:10.3390/molecules22081346 -
Abstract
Adenosine diphosphate-ribose (ADP-ribose) and its derivatives play important roles in a series of complex physiological procedures. The design and synthesis of artificial ADP-ribosylated compounds is an efficient way to develop valuable chemical biology tools and discover new drug candidates. However, the synthesis of
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Adenosine diphosphate-ribose (ADP-ribose) and its derivatives play important roles in a series of complex physiological procedures. The design and synthesis of artificial ADP-ribosylated compounds is an efficient way to develop valuable chemical biology tools and discover new drug candidates. However, the synthesis of ADP-ribosylated compounds is currently difficult due to structural complexity, easily broken pyrophosphate bond and high hydrophilicity. In this paper, ADP-ribosyl-N3 was designed and synthesized for the first time. With ADP-ribosyl-N3 as the key precursor, a divergent post-modification strategy was developed to prepare structurally diverse ADP-ribosylated compounds including novel nucleotides and peptides bearing ADP-ribosyl moieties. Full article
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Open AccessArticle
Two New Tetravacant Organometallic Keggin-Type Heteropolyoxomolybdates-Supported Manganese Carbonyl Derivatives
Molecules 2017, 22(8), 1351; doi:10.3390/molecules22081351 -
Abstract
Two novel heteropolyoxomolybdate [XMo8O31]n− (X = Ge(1) or P(2)) manganese carbonyl derivatives [(CH3)4N]6H6{MnII(GeMo8O31)[MnI(CO)3]2}2
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Two novel heteropolyoxomolybdate [XMo8O31]n− (X = Ge(1) or P(2)) manganese carbonyl derivatives [(CH3)4N]6H6{MnII(GeMo8O31)[MnI(CO)3]2}2·12H2O (1) and [(CH3)4N]4H6{MnII(PMo8O31)[MnI(CO)3]2}2·14H2O (2), have been successfully synthesized and characterized in the solid state by single crystal X-ray diffraction, IR and thermogravimetric analysis, and in solution by UV-Vis spectroscopy and electrochemistry. The two polyoxomolybdate-based organometallic compounds 1 and 2 represent rare examples of transition metal sandwich-based polyoxometalate metal carbonyl derivatives (PMCDs), in which the organic-inorganic hybrids are composed of four Mn(CO)3+ groups symmetrically occupied the tetravacant sites of dimeric heteropolyoxomolybdate {Mn2(XMo8O31)2}n− through MnI-O-Mo bonds. The carbonyl functionalized Mn atoms are octahedrally coordinated via three μ2-oxygens of the [XMo8O31]n− unit and three carbonyl carbon atoms. Interestingly, 1 and 2 form a psedocuboidal ring Mn(CO)3Mo3O12 with {Mn(CO)3}+ occupying the three fold axis of the Mo3O12 octahedral triad. Beside this, the two centrally placed adjacent MnII atoms show intramolecular Mn∙∙∙Mn interactions of 3.11 and 3.16 Å in 1 and 2, respectively. Significant n→π* and O···O intermolecular interactions between the orthogonally aligned adjacent carbonyl groups through the overlap of lone-pair electrons on oxygen atoms with the antibonding orbital (π*) of the adjacent carbony carbon atom of the subsequent units in 1 and 2 were observed. The electrochemical properties of the two compounds were also been investigated. Full article
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Open AccessArticle
Inhibitors of the Detoxifying Enzyme of the Phytoalexin Brassinin Based on Quinoline and Isoquinoline Scaffolds
Molecules 2017, 22(8), 1345; doi:10.3390/molecules22081345 -
Abstract
The detoxification of the phytoalexin brassinin to indole-3-carboxaldehyde and S-methyl dithiocarbamate is catalyzed by brassinin oxidase (BOLm), an inducible fungal enzyme produced by the plant pathogen Leptosphaeria maculans. Twenty-six substituted quinolines and isoquinolines are synthesized and evaluated for antifungal activity against
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The detoxification of the phytoalexin brassinin to indole-3-carboxaldehyde and S-methyl dithiocarbamate is catalyzed by brassinin oxidase (BOLm), an inducible fungal enzyme produced by the plant pathogen Leptosphaeria maculans. Twenty-six substituted quinolines and isoquinolines are synthesized and evaluated for antifungal activity against L. maculans and inhibition of BOLm. Eleven compounds that inhibit BOLm activity are reported, of which 3-ethyl-6-phenylquinoline displays the highest inhibitory effect. In general, substituted 3-phenylquinolines show significantly higher inhibitory activities than the corresponding 2-phenylquinolines. Overall, these results indicate that the quinoline scaffold is a good lead to design paldoxins (phytoalexin detoxification inhibitors) that inhibit the detoxification of brassinin by L. maculans. Full article
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