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1

Chitin biosynthesis enhancement by the endochitinase inhibitor allosamidin.  

PubMed

Membrane preparations of Artemia salina synthetize radiolabelled chitin from UDP-[U-14C]GlcNAc at a low rate (Horst, M.N. (1981) J. Biol. Chem. 256, 1412-1419). We now report that, when the specific endochitinase inhibitor allosamidin is present in addition to the established activators trypsin and GlcNAc, incorporation of [U-14C]GlcNAc into chitin is increased up to 58-fold over the basic synthesis rate. Thus, a greatly enhanced apparent chitin synthase activity is observed in membranes from an arthropod species when simultaneous degradation of chitin is inhibited. PMID:2143906

Peter, M G; Schweikart, F

1990-06-01

2

High resolution genetic mapping uncovers chitin synthase-1 as the target-site of the structurally diverse mite growth inhibitors clofentezine, hexythiazox and etoxazole in Tetranychus urticae.  

PubMed

The acaricides clofentezine, hexythiazox and etoxazole are commonly referred to as 'mite growth inhibitors', and clofentezine and hexythiazox have been used successfully for the integrated control of plant mite pests for decades. Although they are still important today, their mode of action has remained elusive. Recently, a mutation in chitin synthase 1 (CHS1) was linked to etoxazole resistance. In this study, we identified and investigated a Tetranychus urticae strain (HexR) harboring recessive, monogenic resistance to each of hexythiazox, clofentezine, and etoxazole. To elucidate if there is a common genetic basis for the observed cross-resistance, we adapted a previously developed bulk segregant analysis method to map with high resolution a single, shared resistance locus for all three compounds. This finding indicates that the underlying molecular basis for resistance to all three compounds is identical. This locus is centered on the CHS1 gene, and as supported by additional genetic and biochemical studies, a non-synonymous variant (I1017F) in CHS1 associates with resistance to each of the tested acaricides in HexR. Our findings thus demonstrate a shared molecular mode of action for the chemically diverse mite growth inhibitors clofentezine, hexythiazox and etoxazole as inhibitors of an essential, non-catalytic activity of CHS1. Given the previously documented cross-resistance between clofentezine, hexythiazox and the benzyolphenylurea (BPU) compounds flufenoxuron and cycloxuron, CHS1 should be also considered as a potential target-site of insecticidal BPUs. PMID:24859419

Demaeght, Peter; Osborne, Edward J; Odman-Naresh, Jothini; Grbi?, Miodrag; Nauen, Ralf; Merzendorfer, Hans; Clark, Richard M; Van Leeuwen, Thomas

2014-08-01

3

2-acylamido analogues of N-acetylglucosamine prime formation of chitin oligosaccharides by yeast chitin synthase 2  

Technology Transfer Automated Retrieval System (TEKTRAN)

Chitin, a polymer of beta-1,4-linked N-acetylglucosamine (GlcNAc), is a key component of the cell walls of fungi and the exoskeletons of arthropods. Chitin synthases (CSs) transfer GlcNAc from UDP-GlcNAc to pre-existing chitin chains in reactions that are typically stimulated by free GlcNAc. The eff...

4

Characterization of a Chitin Synthase Encoding Gene and Effect of Diflubenzuron in Soybean Aphid, Aphis Glycines  

PubMed Central

Chitin synthases are critical enzymes for synthesis of chitin and thus for subsequent growth and development in insects. We identified the cDNA of chitin synthase gene (CHS) in Aphis glycines, the soybean aphid, which is a serious pest of soybean. The full-length cDNA of CHS in A. glycines (AyCHS) was 5802 bp long with an open reading frame of 4704 bp that encoded for a 1567 amino acid residues protein. The predicted AyCHS protein had a molecular mass of 180.05 kDa and its amino acid sequence contained all the signature motifs (EDR, QRRRW and TWGTR) of chitin synthases. The quantitative real-time PCR (qPCR) analysis revealed that AyCHS was expressed in all major tissues (gut, fat body and integument); however, it had the highest expression in integument (~3.5 fold compared to gut). Interestingly, the expression of AyCHS in developing embryos was nearly 7 fold higher compared to adult integument, which probably is a reflection of embryonic molts in hemimetabolus insects. Expression analysis in different developmental stages of A. glycines revealed a consistent AyCHS expression in all stages. Further, through leaf dip bioassay, we tested the effect of diflubenzuron (DFB, Dimilin ®), a chitin-synthesis inhibitor, on A. glycines' survival, fecundity and body weight. When fed with soybean leaves previously dipped in 50 ppm DFB solution, A. glycines nymphs suffered significantly higher mortality compared to control. A. glycines nymphs feeding on diflubenzuron treated leaves showed a slightly enhanced expression (1.67 fold) of AyCHS compared to nymphs on untreated leaves. We discussed the potential applications of the current study to develop novel management strategies using chitin-synthesis inhibitors and using RNAi by knocking down AyCHS expression. PMID:23139631

Bansal, Raman; Mian, M. A. Rouf; Mittapalli, Omprakash; Michel, Andy P.

2012-01-01

5

Chitin synthases are required for survival, fecundity and egg-hatch in the red flour beetle, Tribolium castaneum  

Technology Transfer Automated Retrieval System (TEKTRAN)

The synthesis of chitin, the Beta-1,4-linked polymer of N-acetylglucosamine, is catalyzed by chitin synthase (CHS). Chitin is essential for the structural integrity of the exoskeletal cuticle and midgut peritrophic membrane (PM) of insects. To study the functions of the two chitin synthase genes, ...

6

CAL1, a gene required for activity of chitin synthase 3 in Saccharomyces cerevisiae  

PubMed Central

The CAL1 gene was cloned by complementation of the defect in Calcofluor- resistant calR1 mutants of Saccharomyces cerevisiae. Transformation of the mutants with a plasmid carrying the appropriate insert restored Calcofluor sensitivity, wild-type chitin levels and normal spore maturation. Southern blots using the DNA fragment as a probe showed hybridization to a single locus. Allelic tests indicated that the cloned gene corresponded to the calR1 locus. The DNA insert contains a single open-reading frame encoding a protein of 1,099 amino acids with a molecular mass of 124 kD. The predicted amino acid sequence shows several regions of homology with those of chitin synthases 1 and 2 from S. cerevisiae and chitin synthase 1 from Candida albicans. calR1 mutants have been found to be defective in chitin synthase 3, a trypsin- independent synthase. Transformation of the mutants with a plasmid carrying CAL1 restored chitin synthase 3 activity; however, overexpression of the enzyme was not achieved even with a high copy number plasmid. Since Calcofluor-resistance mutations different from calR1 also result in reduced levels of chitin synthase 3, it is postulated that the products of some of these CAL genes may be limiting for expression of the enzymatic activity. Disruption of the CAL1 gene was not lethal, indicating that chitin synthase 3 is not an essential enzyme for S. cerevisiae. PMID:2050737

1991-01-01

7

Evolution and phylogenetic relationships of chitin synthases from yeasts and fungi  

Microsoft Academic Search

Chitin, the structural component that provides rigidity to the cell wall of fungi is the product of chitin synthases (Chs). These enzymes are not restricted to fungi, but are amply distributed in four of the five eukaryotic ‘crown kingdoms’. Dendrograms obtained by multiple alignment of Chs revealed that fungal enzymes can be classified into two divisions that branch into at

José Ruiz-Herrera; Juan Manuel González-Prieto; Roberto Ruiz-Medrano

2002-01-01

8

Sequence of a cDNA and expression of the gene encoding a putative epidermal chitin synthase of Manduca sexta  

Microsoft Academic Search

Glycosyltransferases are enzymes that synthesize oligosaccharides, polysaccharides and glycoconjugates. One type of glycosyltransferase is chitin synthase, a very important enzyme in biology, which is utilized by insects, fungi, and other invertebrates to produce chitin, a polysaccharide of ?-1,4-linked N-acetylglucosamine. Chitin is an important component of the insect’s exoskeletal cuticle and gut lining. To identify and characterize a chitin synthase gene

Yu-Cheng Zhu; Charles A. Specht; Neal T. Dittmer; Subbaratnam Muthukrishnan; Michael R. Kanost; Karl J. Kramer

2002-01-01

9

Early Divergence, Broad Distribution, and High Diversity of Animal Chitin Synthases  

PubMed Central

Even though chitin is one of the most abundant biopolymers in nature, current knowledge on chitin formation is largely based only on data from fungi and insects. This study reveals unanticipated broad taxonomic distribution and extensive diversification of chitin synthases (CSs) in Metazoa, shedding new light on the relevance of chitin in animals and suggesting unforeseen complexity of chitin synthesis in many groups. We uncovered robust orthologs to insect type CSs in several representatives of deuterostomes, which generally are not thought to possess chitin. This suggests a broader distribution and function of chitin in this branch of the animal kingdom. We characterize a new CS type present not only in basal metazoans such as sponges and cnidarians but also in several bilaterian representatives. The most extensive diversification of CSs took place during emergence of lophotrochozoans, the third large group of protostomes next to arthropods and nematodes, resulting in coexistence of up to ten CS paralogs in molluscs. Independent fusion to different kinds of myosin motor domains in fungi and lophotrochozoans points toward high relevance of CS interaction with the cytoskeleton for fine-tuned chitin secretion. Given the fundamental role that chitin plays in the morphology of many animals, the here presented CS diversification reveals many evolutionary complexities. Our findings strongly suggest a very broad and multifarious occurrence of chitin and question an ancestral role as cuticular component. The molecular mechanisms underlying regulation of animal chitin synthesis are most likely far more complex and diverse than existing data from insects suggest. PMID:24443419

Zakrzewski, Anne-C.; Weigert, Anne; Helm, Conrad; Adamski, Marcin; Adamska, Maja; Bleidorn, Christoph; Raible, Florian; Hausen, Harald

2014-01-01

10

Biochemical characterization of chitin synthase activity and inhibition in the African malaria mosquito, Anopheles gambiae  

PubMed Central

Chitin synthase (CHS) is an important enzyme catalyzing the formation of chitin polymers in all chitin containing organisms and a potential target site for insect pest control. However, our understanding of biochemical properties of insect chitin synthases has been very limited. We here report enzymatic and inhibitory properties of chitin synthase prepared from the African malaria mosquito, Anopheles gambiae. Our study, which represents the first time to use a nonradioactive method to assay chitin synthase activity in an insect species, determined the optimal conditions for measuring the enzyme activity, including pH, temperature, and concentrations of the substrate UDP–N-acetyl-D-glucosamine (GlcNAc) and Mg++. The optimal pH was about 6.5–7.0, and the highest activity was detected at temperatures between 37 and 44°C. Dithithreitol is required to prevent melanization of the enzyme extract. CHS activity was enhanced at low concentration of GlcNAc, but inhibited at high concentrations. Proteolytic activation of the activity is significant both in the 500×g supernatant and the 40,000×g pellet. Our study revealed only slight in vitro inhibition of An. gambiae CHS activity by diflubenzuron and nikkomycin Z at the highest concentration (2.5 ?mol/L) examined. There was no in vitro inhibition by polyoxin D at any concentration examined. Furthermore, we did not observe any in vivo inhibition of chitin synthase activity by any of these chemicals at any concentration examined. Our results suggest that the inhibition of chitin synthesis by these chemicals is not due to direct inhibition of chitin synthase in An. gambiae. PMID:23955856

Zhang, Xin; Zhu, Kun Yan

2012-01-01

11

Inhibitors of the fungal cell wall. Synthesis of 4-aryl-4-N-arylamine-1-butenes and related compounds with inhibitory activities on beta(1-3) glucan and chitin synthases.  

PubMed

As part of our project devoted to the search for antifungal agents, which act via a selective mode of action, we synthesized a series of new 4-aryl- or 4-alkyl-N-arylamine-1-butenes and transformed some of them into 2-substituted 4-methyl-tetrahydroquinolines and quinolines by using a novel three-step synthesis. Results obtained in agar dilution assays have shown that 4-aryl homoallylamines not possessing halogen in their structures, tetrahydroquinolines and quinolines, display a range of antifungal properties in particular against Epidermophyton floccosum and Microsporum canis. Regarding the mode of action, all active compounds showed in vitro inhibitory activities against beta(1-3) glucan-synthase and mainly against chitin-synthase. These enzymes catalyze the synthesis of beta(1-3) glucan and chitin, respectively, major polymers of the fungal cell wall. Since fungal but not mammalian cells are encased in a cell wall, its inhibition may represent a useful mode of action for these antifungal compounds. PMID:10819157

Urbina, J M; Cortés, J C; Palma, A; López, S N; Zacchino, S A; Enriz, R D; Ribas, J C; Kouznetzov, V V

2000-04-01

12

Deletion of a Chitin Synthase Gene in a Citric Acid Producing Strain of Aspergillus niger  

SciTech Connect

Citric acid production by the filamentous fungus Aspergillus niger is carried out in a process that causes the organism to drastically alter its morphology. This altered morphology includes hyphal swelling and highly limited polar growth resulting in clumps of swollen cells that eventually aggregate into pellets of approximately 100 microns in diameter. In this pelleted form, A. niger has increased citric acid production as compared to growth in filamentous form. Chitin is a crucial component of the cell wall of filamentous fungi. Alterations in the deposition or production of chitin may have profound effects on the morphology of the organism. In order to study the role of chitin synthesis in pellet formation we have deleted a chitin synthase gene (csmA) in Aspergillus niger strain ATCC 11414 using a PCR based deletion construct. This class of chitin synthases is only found in filamentous fungi and is not present in yeasts. The csmA genes contain a myosin motor domain at the N-terminus and a chitin synthesis domain at the C-terminus. They are believed to contribute to the specialized polar growth observed in filamentous fungi that is lacking in yeasts. The csmA deletion strain (csmA?) was subjected to minimal media with and without osmotic stabilizers as well as tested in citric acid production media. Without osmotic stabilizers, the mutant germlings were abnormally swollen, primarily in the subapical regions, and contained large vacuoles. However, this swelling is ultimately not inhibitory to growth as the germlings are able to recover and undergo polar growth. Colony formation was largely unaffected in the absence of osmotic stabilizers. In citric acid production media csmA? was observed to have a 2.5 fold increase in citric acid production. The controlled expression of this class of chitin synthases may be useful for improving production of organic acids in filamentous fungi.

Rinker, Torri E.; Baker, Scott E.

2007-01-29

13

Identification of the conserved coding sequences of three chitin synthase genes in Fonsecaea pedrosoi.  

PubMed

Primers having designs based on highly conserved stretches in the deduced amino acid sequences of chitin synthase (CHS) genes were used in PCR reactions to amplify 600 bp and 366 bp products from the genomic DNA of three major causal agents of chromoblastomycosis. Cloning and sequencing of the PCR products of one of these fungi, Fonsecaea pedrosoi, identified three CHS sequences designated as FpCHS1, FpCHS2 and FpCHS3. FpCHS1 and FpCHS2 were homologous to regions of CHS1 and CHS2 of Saccharomyces cerevisiae, and their derived amino acid sequences fell into chitin synthase classes I and II, respectively. FpCHS3 was homologous to a region of the CAL1/CSD2 gene of S. cerevisiae, which codes for the chitin synthase three (Chs3) enzyme in that fungus. Phylogenetic trees constructed using the deduced amino acid sequences of PCR-amplified CHS products from many fungi clustered F. pedrosoi with other dematiaceous fungi, providing new molecular evidence for the genetic relatedness of these organisms. The identification of these CHS genes in F. pedrosoi will facilitate future studies of the functional roles of chitin synthases in the unique in vivo dimorphism exhibited by chromoblastomycotic fungi. PMID:8732357

Karuppayil, S M; Peng, M; Mendoza, L; Levins, T A; Szaniszlo, P J

1996-01-01

14

Identification and characterization of two chitin synthase genes in African malaria mosquito, Anopheles gambiae  

PubMed Central

Chitin synthase (CHS) represents an attractive target site for combating insect pests as insect growth and development are strictly dependent on precisely tuned chitin biosynthesis and this pathway is absent in humans and other vertebrates. Current knowledge on CHS in insects, especially their structures, functions, and regulations is still very limited. We report the identification and characterization of two chitin synthase genes, AgCHS1 and AgCHS2, in African malaria mosquito, Anopheles gambiae. AgCHS1 and AgCHS2 were predicted to encode proteins of 1,578 and 1,586 amino acid residues, respectively. Their deduced amino acid sequences show high similarities to other insect chitin synthases. Transcriptional analysis indicated that AgCHS1 was expressed in egg, larval, pupal and adult stages whereas AgCHS2 appeared to be expressed at relatively low levels, particularly during the larval stages as examined by reverse transcription (RT)-PCR and real-time quantitative PCR. Relatively high expression was detected in the carcass followed by the foregut and hindgut for AgCHS1, and the foregut (cardia included) followed by the midgut for AgCHS2. Fluorescence in situ hybridization (FISH) and immunohistochemical analysis revealed new information including the localization of the two enzymes in the ommatidia of the compound eyes, and AgCHS2 in the thoracic and abdominal inter-segmental regions of pupal integument. PMID:22683441

Zhang, Xin; Zhang, Jianzhen; Park, Yoonseong; Zhu, Kun Yan

2012-01-01

15

WdChs4p, a Homolog of Chitin Synthase 3 in Saccharomyces cerevisiae, Alone Cannot Support Growth of Wangiella (Exophiala) dermatitidis at the Temperature of Infection  

Microsoft Academic Search

By using improved transformation methods for Wangiella dermatitidis, and a cloned fragment of its chitin synthase 4 structural gene (WdCHS4) as a marking sequence, the full-length gene was rescued from the genome of this human pathogenic fungus. The encoded chitin synthase product (WdChs4p) showed high homology with Chs3p of Saccharomyces cerevisiae and other class IV chitin synthases, and Northern blotting

ZHENG WANG; LI ZHENG; MELINDA HAUSER; JEFFERY M. BECKER; PAUL J. SZANISZLO

1999-01-01

16

Exposure to Diflubenzuron Results in an Up-Regulation of a Chitin Synthase 1 Gene in Citrus Red Mite, Panonychus citri (Acari: Tetranychidae)  

PubMed Central

Chitin synthase synthesizes chitin, which is critical for the arthropod exoskeleton. In this study, we cloned the cDNA sequences of a chitin synthase 1 gene, PcCHS1, in the citrus red mite, Panonychus citri (McGregor), which is one of the most economically important pests of citrus worldwide. The full-length cDNA of PcCHS1 contains an open reading frame of 4605 bp of nucleotides, which encodes a protein of 1535 amino acid residues with a predicted molecular mass of 175.0 kDa. A phylogenetic analysis showed that PcCHS1 was most closely related to CHS1 from Tetranychus urticae. During P. citri development, PcCHS1 was constantly expressed in all stages but highly expressed in the egg stage (114.8-fold higher than in the adult). When larvae were exposed to diflubenzuron (DFB) for 6 h, the mite had a significantly high mortality rate, and the mRNA expression levels of PcCHS1 were significantly enhanced. These results indicate a promising use of DFB to control P. citri, by possibly acting as an inhibitor in chitin synthesis as indicated by the up-regulation of PcCHS1 after exposure to DFB. PMID:24590130

Xia, Wen-Kai; Ding, Tian-Bo; Niu, Jin-Zhi; Liao, Chong-Yu; Zhong, Rui; Yang, Wen-Jia; Liu, Bin; Dou, Wei; Wang, Jin-Jun

2014-01-01

17

Bait Matrix for Delivery of Chitin Synthesis Inhibitors to the Formosan Subterranean Termite (Isoptera: Rhinotermitidae)  

Microsoft Academic Search

The efficacy of three chitin synthesis inhibitors, diflubenzuron, hexaflumuron, and chlorfluazuron, incorporated into a novel bait matrix to kill the Formosan subterranean termite, Coptotermes formosanus Shiraki, was evaluated in the laboratory. The bait matrix was significantly preferred by C. formosanus over southern yellow pine wood in a two-choice feeding test. Bait formulations containing 250 ppm of the three chitin synthesis

M. Guadalupe Rojas; J. A. Morales-Ramos

2001-01-01

18

Different Chitin Synthase Genes Are Required for Various Developmental and Plant Infection Processes in the Rice Blast Fungus Magnaporthe oryzae  

PubMed Central

Chitin is a major component of fungal cell wall and is synthesized by chitin synthases (Chs). Plant pathogenic fungi normally have multiple chitin synthase genes. To determine their roles in development and pathogenesis, we functionally characterized all seven CHS genes in Magnaporthe oryzae. Three of them, CHS1, CHS6, and CHS7, were found to be important for plant infection. While the chs6 mutant was non-pathogenic, the chs1 and chs7 mutants were significantly reduced in virulence. CHS1 plays a specific role in conidiogenesis, an essential step for natural infection cycle. Most of chs1 conidia had no septum and spore tip mucilage. The chs6 mutant was reduced in hyphal growth and conidiation. It failed to penetrate and grow invasively in plant cells. The two MMD-containing chitin synthase genes, CHS5 and CHS6, have a similar expression pattern. Although deletion of CHS5 had no detectable phenotype, the chs5 chs6 double mutant had more severe defects than the chs6 mutant, indicating that they may have overlapping functions in maintaining polarized growth in vegetative and invasive hyphae. Unlike the other CHS genes, CHS7 has a unique function in appressorium formation. Although it was blocked in appressorium formation by germ tubes on artificial hydrophobic surfaces, the chs7 mutant still produced melanized appressoria by hyphal tips or on plant surfaces, indicating that chitin synthase genes have distinct impacts on appressorium formation by hyphal tip and germ tube. The chs7 mutant also was defective in appressorium penetration and invasive growth. Overall, our results indicate that individual CHS genes play diverse roles in hyphal growth, conidiogenesis, appressorium development, and pathogenesis in M. oryzae, and provided potential new leads in the control of this devastating pathogen by targeting specific chitin synthases. PMID:22346755

Li, Guo-Tian; Qi, Lin-Lu; Zhang, Yu-Jun; Wang, Chen-Fang; Zhao, Wen-Sheng; Xu, Jin-Rong; Peng, You-Liang

2012-01-01

19

Fusarium verticillioides chitin synthases CHS5 and CHS7 are required for normal growth and pathogenicity.  

PubMed

Fusarium verticillioides is both an endophyte and a pathogen of maize and is a health threat in many areas of the world because it can contaminate maize with fumonisins, a toxic secondary metabolite. We identified eight putative chitin synthase (CHS) genes in F. verticillioides genomic sequence, and phylogenetic evidence shows that they group into seven established CHS gene classes. We targeted two CHSs (CHS5 and CHS7) for deletion analysis and found that both are required for normal hyphal growth and maximal disease of maize seedlings and ears. CHS5 and CHS7 encode a putative class V and class VII fungal chitin synthase, respectively; they are located adjacent to each other and are divergently transcribed. Fluorescent microscopy found that both CHS deficient strains produce balloon-shaped hyphae, while growth assays indicated that they were more sensitive to cell wall stressing compounds (e.g., the antifungal compound Nikkomycin Z) than wild type. Pathogenicity assays on maize seedlings and ears indicated that both strains were significantly reduced in their ability to cause disease. Our results demonstrate that both CHS5 and CHS7 are necessary for proper hyphal growth and pathogenicity of F. verticillioides on maize. PMID:21246198

Larson, Troy M; Kendra, David F; Busman, Mark; Brown, Daren W

2011-06-01

20

Physiological and Morphological Aspects of Aedes aegypti Developing Larvae: Effects of the Chitin Synthesis Inhibitor Novaluron  

PubMed Central

Population control of the dengue vector mosquito, Aedes aegypti, is difficult due to many reasons, one being the development of resistance to neurotoxic insecticides employed. The biosynthesis of chitin, a major constituent of insect cuticle, is a novel target for population control. Novaluron is a benzoylphenylurea (BPU) that acts as a chitin synthesis inhibitor, already used against mosquitoes. However, information regarding BPU effects on immature mosquito stages and physiological parameters related with mosquito larval development are scarce. A set of physiological parameters were recorded in control developing larvae and novaluron was administered continuously to Ae. aegypti larvae, since early third instar. Larval instar period duration was recorded from third instar until pupation. Chitin content was measured during third and fourth instars. Fourth instars were processed histochemically at the mesothorax region, stained with hematoxylin and eosin (HE) for assessment of internal tissues, and labeled with WGA-FITC to reveal chitinized structures. In control larvae: i) there is a chitin content increase during both third and fourth instars where late third instars contain more chitin than early fourth instars; ii) thoracic organs and a continuous cuticle, closely associated with the underlying epidermis were observed; iii) chitin was continuously present throughout integument cuticle. Novaluron treatment inhibited adult emergence, induced immature mortality, altered adult sex ratio and caused delay in larval development. Moreover, novaluron: i) significantly affected chitin content during larval development; ii) induced a discontinuous and altered cuticle in some regions while epidermis was often thinner or missing; iii) rendered chitin cuticle presence discontinuous and less evident. In both control and novaluron larvae, chitin was present in the peritrophic matrix. This study showed quantitatively and qualitatively evidences of novaluron effects on Ae. aegypti larval development. To our knowledge, this is the first report describing histological alterations produced by a BPU in immature vector mosquitoes. PMID:22291942

Farnesi, Luana C.; Brito, José M.; Linss, Jutta G.; Pelajo-Machado, Marcelo; Valle, Denise; Rezende, Gustavo L.

2012-01-01

21

Cloning and sequence analysis of chitin synthase gene fragments of Demodex mites.  

PubMed

To our knowledge, few reports on Demodex studied at the molecular level are available at present. In this study our group, for the first time, cloned, sequenced and analyzed the chitin synthase (CHS) gene fragments of Demodex folliculorum, Demodex brevis, and Demodex canis (three isolates from each species) from Xi'an China, by designing specific primers based on the only partial sequence of the CHS gene of D. canis from Japan, retrieved from GenBank. Results show that amplification was successful only in three D. canis isolates and one D. brevis isolate out of the nine Demodex isolates. The obtained fragments were sequenced to be 339 bp for D. canis and 338 bp for D. brevis. The CHS gene sequence similarities between the three Xi'an D. canis isolates and one Japanese D. canis isolate ranged from 99.7% to 100.0%, and those between four D. canis isolates and one D. brevis isolate were 99.1%-99.4%. Phylogenetic trees based on maximum parsimony (MP) and maximum likelihood (ML) methods shared the same clusters, according with the traditional classification. Two open reading frames (ORFs) were identified in each CHS gene sequenced, and their corresponding amino acid sequences were located at the catalytic domain. The relatively conserved sequences could be deduced to be a CHS class A gene, which is associated with chitin synthesis in the integument of Demodex mites. PMID:23024043

Zhao, Ya-e; Wang, Zheng-hang; Xu, Yang; Xu, Ji-ru; Liu, Wen-yan; Wei, Meng; Wang, Chu-ying

2012-10-01

22

Cloning and sequence analysis of chitin synthase gene fragments of Demodex mites*  

PubMed Central

To our knowledge, few reports on Demodex studied at the molecular level are available at present. In this study our group, for the first time, cloned, sequenced and analyzed the chitin synthase (CHS) gene fragments of Demodex folliculorum, Demodex brevis, and Demodex canis (three isolates from each species) from Xi’an China, by designing specific primers based on the only partial sequence of the CHS gene of D. canis from Japan, retrieved from GenBank. Results show that amplification was successful only in three D. canis isolates and one D. brevis isolate out of the nine Demodex isolates. The obtained fragments were sequenced to be 339 bp for D. canis and 338 bp for D. brevis. The CHS gene sequence similarities between the three Xi’an D. canis isolates and one Japanese D. canis isolate ranged from 99.7% to 100.0%, and those between four D. canis isolates and one D. brevis isolate were 99.1%–99.4%. Phylogenetic trees based on maximum parsimony (MP) and maximum likelihood (ML) methods shared the same clusters, according with the traditional classification. Two open reading frames (ORFs) were identified in each CHS gene sequenced, and their corresponding amino acid sequences were located at the catalytic domain. The relatively conserved sequences could be deduced to be a CHS class A gene, which is associated with chitin synthesis in the integument of Demodex mites. PMID:23024043

Zhao, Ya-e; Wang, Zheng-hang; Xu, Yang; Xu, Ji-ru; Liu, Wen-yan; Wei, Meng; Wang, Chu-ying

2012-01-01

23

THE EFFECT OF AN INSECT CHITIN SYNTHESIS INHIBITOR ON HONEY BEES  

E-print Network

THE EFFECT OF AN INSECT CHITIN SYNTHESIS INHIBITOR ON HONEY BEES E.W. HERBERT, Jr. R.J. ARGAUER * H. SHIMANUKI * U.S. Department of Agriculture, Agricultural Research Service """ Bioenvironmental Bee-flying colonies of honey bees. Brood rearing was temporarily terminated for a period of 2-3 weeks depending

Paris-Sud XI, Université de

24

The Myosin Motor Domain of Fungal Chitin Synthase V Is Dispensable for Vesicle Motility but Required for Virulence of the Maize Pathogen Ustilago maydis[W  

PubMed Central

Class V chitin synthases are fungal virulence factors required for plant infection. They consist of a myosin motor domain fused to a membrane-spanning chitin synthase region that participates in fungal cell wall formation. The function of the motor domain is unknown, but it might deliver the myosin chitin synthase-attached vesicles to the growth region. Here, we analyze the importance of both domains in Mcs1, the chitin synthase V of the maize smut fungus Ustilago maydis. By quantitative analysis of disease symptoms, tissue colonization, and single-cell morphogenic parameters, we demonstrate that both domains are required for fungal virulence. Fungi carrying mutations in the chitin synthase domain are rapidly recognized and killed by the plant, whereas fungi carrying a deletion of the motor domain show alterations in cell wall composition but can invade host tissue and cause a moderate plant response. We also show that Mcs1-bound vesicles exhibit long-range movement for up to 20 ?m at a velocity of ~1.75 ?m/s. Apical Mcs1 localization depends on F-actin and the motor domain, whereas Mcs1 motility requires microtubules and persists when the Mcs1 motor domain is deleted. Our results suggest that the myosin motor domain of ChsV supports exocytosis but not long-range delivery of transport vesicles. PMID:20663961

Treitschke, Steffi; Doehlemann, Gunther; Schuster, Martin; Steinberg, Gero

2010-01-01

25

Nitric oxide synthase inhibitors and cerebral vasospasm.  

PubMed

L-arginine is a source of nitric oxide (NO) that is cleaved from the terminal guanidino nitrogen atom by nitric oxide synthase (NOS). NO evokes, because of its free radical properties and affinity to heme, ferrous iron and cysteine, a wide spectrum of physiological and pathophysiological effects. For many years, different exogenous NOS inhibitors were used to elucidate the role of NOS and NO in health and disease. Later, endogenous NOS inhibitors, as asymmetric dimethylarginine (ADMA) were discovered. Endogenous inhibitors as ADMA are produced by post-translational methylation of L-arginine which is catalyzed by a family of protein N-methyltransferases (PRMT), using S-adenosylmethionine as a methyl group donor. ADMA is eliminated by dimethylarginine dimethylaminohydrolases (DDAH I or II). ADMA hydrolysis increases NOS activity and NO production. Furthermore, L-citrulline, a by-product of ADMA hydrolysis as well as of NO production by NOS, can in turn inhibit DDAH. Therefore, endogenous inhibition of NOS can be modified via different ways (1) changing the availability of L-arginine and/or of L-citrulline; (2) stimulating or inhibiting DDAH activity; (3) modifying methylation via regulating availability of adenosylmethionine; or (4) modifying PRMT activity. Research elucidating the role of NOS inhibitors in respect of delayed cerebral vasospasm after subarachnoid hemorrhage is summarized. PMID:21116921

Jung, C S

2011-01-01

26

Molecular Cloning, Characterization and mRNA Expression of a Chitin Synthase 2 Gene from the Oriental Fruit Fly, Bactrocera dorsalis (Diptera: Tephritidae)  

PubMed Central

Chitin synthase (CHS), a potential target for eco-friendly insecticides, plays an essential role in chitin formation in insects. In this study, a full-length cDNA encoding chitin synthase 2 (BdCHS2) was cloned and characterized in the oriental fruit fly, Bactrocera dorsalis. The BdCHS2 cDNA had 4417 nucleotides, containing an open reading frame of 4122 nucleotides, which encoded 1373 amino acid residues with a predicted molecular weight of 158.5 kDa. Phylogenetic analysis with other insect CHSs suggested that BdCHS2 belongs to insect CHS2. The BdCHS2 transcript was predominately found in midgut but was detected at low levels in fat body, Malpighian tubules, integument, and trachea. Moreover, BdCHS2 was expressed in all developmental stages, and highly expressed in the feeding stages. There was a positive relationship between BdCHS2 expression and total chitin content during development. Furthermore, both the gene expression and chitin content in midgut decreased when the insect was fed for 24 h, then starved for 24 h, while they increased dramatically and rapidly under the condition of starvation for 24 h then feeding for 24 h. These results suggest that BdCHS2 may play an important role in regulating chitin content of the midgut, and subsequently affect the growth and development of B. dorsalis. PMID:23965972

Chen, Li; Yang, Wen-Jia; Cong, Lin; Xu, Kang-Kang; Wang, Jin-Jun

2013-01-01

27

Large-Scale Phylogenetic Classification of Fungal Chitin Synthases and Identification of a Putative Cell-Wall Metabolism Gene Cluster in Aspergillus Genomes  

PubMed Central

The cell wall is a protective and versatile structure distributed in all fungi. The component responsible for its rigidity is chitin, a product of chitin synthase (Chsp) enzymes. There are seven classes of chitin synthase genes (CHS) and the amount and type encoded in fungal genomes varies considerably from one species to another. Previous Chsp sequence analyses focused on their study as individual units, regardless of genomic context. The identification of blocks of conserved genes between genomes can provide important clues about the interactions and localization of chitin synthases. On the present study, we carried out an in silico search of all putative Chsp encoded in 54 full fungal genomes, encompassing 21 orders from five phyla. Phylogenetic studies of these Chsp were able to confidently classify 347 out of the 369 Chsp identified (94%). Patterns in the distribution of Chsp related to taxonomy were identified, the most prominent being related to the type of fungal growth. More importantly, a synteny analysis for genomic blocks centered on class IV Chsp (the most abundant and widely distributed Chsp class) identified a putative cell wall metabolism gene cluster in members of the genus Aspergillus, the first such association reported for any fungal genome. PMID:25148134

Pacheco-Arjona, Jose Ramon; Ramirez-Prado, Jorge Humberto

2014-01-01

28

A cellular model for screening neuronal nitric oxide synthase inhibitors  

Microsoft Academic Search

Nitric oxide synthase (NOS) inhibitors are potential drug candidates because it has been well demonstrated that excessive production of nitric oxide critically contributes to a range of diseases. Most inhibitors have been screened in vitro using recombinant enzymes, leading to the discovery of a variety of potent compounds. To make inhibition studies more physiologically relevant and bridge the gap between

Jianguo Fang; Richard B. Silverman

2009-01-01

29

Farnesyl Diphosphate Synthase Inhibitors from In Silico Screening  

PubMed Central

The relaxed complex scheme is an in silico drug screening method that accounts for receptor flexibility using molecular dynamics simulations. Here, we used this approach combined with similarity searches and experimental inhibition assays to identify several low micromolar, non-bisphosphonate inhibitors, bisamidines, of farnesyl diphosphate synthase (FPPS), an enzyme targeted by some anticancer and antimicrobial agents and for the treatment of bone resorption diseases. This novel class of farnesyl diphosphate synthase inhibitors have more drug-like properties than existing bisphosphonate inhibitors, making them interesting pharmaceutical leads. PMID:23421555

Lindert, Steffen; Zhu, Wei; Liu, Yi-Liang; Pang, Ran; Oldfield, Eric; McCammon, J Andrew

2013-01-01

30

A Structural and Biochemical Model of Processive Chitin Synthesis*  

PubMed Central

Chitin synthases (CHS) produce chitin, an essential component of the fungal cell wall. The molecular mechanism of processive chitin synthesis is not understood, limiting the discovery of new inhibitors of this enzyme class. We identified the bacterial glycosyltransferase NodC as an appropriate model system to study the general structure and reaction mechanism of CHS. A high throughput screening-compatible novel assay demonstrates that a known inhibitor of fungal CHS also inhibit NodC. A structural model of NodC, on the basis of the recently published BcsA cellulose synthase structure, enabled probing of the catalytic mechanism by mutagenesis, demonstrating the essential roles of the DD and QXXRW catalytic motifs. The NodC membrane topology was mapped, validating the structural model. Together, these approaches give insight into the CHS structure and mechanism and provide a platform for the discovery of inhibitors for this antifungal target. PMID:24942743

Dorfmueller, Helge C.; Ferenbach, Andrew T.; Borodkin, Vladimir S.; van Aalten, Daan M. F.

2014-01-01

31

A structural and biochemical model of processive chitin synthesis.  

PubMed

Chitin synthases (CHS) produce chitin, an essential component of the fungal cell wall. The molecular mechanism of processive chitin synthesis is not understood, limiting the discovery of new inhibitors of this enzyme class. We identified the bacterial glycosyltransferase NodC as an appropriate model system to study the general structure and reaction mechanism of CHS. A high throughput screening-compatible novel assay demonstrates that a known inhibitor of fungal CHS also inhibit NodC. A structural model of NodC, on the basis of the recently published BcsA cellulose synthase structure, enabled probing of the catalytic mechanism by mutagenesis, demonstrating the essential roles of the DD and QXXRW catalytic motifs. The NodC membrane topology was mapped, validating the structural model. Together, these approaches give insight into the CHS structure and mechanism and provide a platform for the discovery of inhibitors for this antifungal target. PMID:24942743

Dorfmueller, Helge C; Ferenbach, Andrew T; Borodkin, Vladimir S; van Aalten, Daan M F

2014-08-15

32

Characterization of microsomal prostaglandin E synthase 1 inhibitors.  

PubMed

Microsomal prostaglandin E synthase-1 (mPGES-1) is an inducible terminal synthase in PGE2 biosynthesis by inflammatory and cancer cells. Clinical and experimental data emphasize that mPGES-1 might be a valuable target, with improved selectivity and safety compared to traditional NSAIDs or selective COX-2 inhibitors, in the treatment of inflammatory diseases, different types of cancer as well as central symptoms elicited by peripheral inflammation. Since the first characterization of mPGES-1, the numbers of publications on mPGES-1 structure, pathogenic role and inhibitor development have increased exponentially; however, there are currently no selective mPGES-1 inhibitors available for clinical use. In this MiniReview, we focus on recent advances in the development of selective inhibitors of mPGES-1 activity, with the aim to discuss the effects of targeting mPGES-1 in different inflammatory models in vitro and in vivo. PMID:24138533

Korotkova, Marina; Jakobsson, Per-Johan

2014-01-01

33

Effects of four chitin synthesis inhibitors on feeding and mortality of the Eastern subterranean termite, Reticulitermes flavipes Kollar (Isoptera:Rhinotermitidae)  

E-print Network

This study measured changes in feeding and mortality of the Eastern subterranean termite, Reticulitermes flavipes when exposed to diets treated with one of four chitin synthesis inhibitors including; diflubenzuron, triflumuron, hexaflumuron...

Vahabzadeh, Rebecca D.

2012-06-07

34

Evaluation of Two Formulated Chitin Synthesis Inhibitors, Hexaflumuron and Lufenuron Against the Raisin Moth, Ephestia figulilella  

PubMed Central

The raisin moth, Ephestia figulilella Gregson (Lepidoptera: Pyralidae), has a nearly cosmopolitan distribution, and causes severe quantitative and qualitative losses throughout the world. The larvae attack various drying and dried fruits, fallen figs, and damaged or moldy clusters of grapes on vines. Control of this pest in storage depends mostly on synthetic pesticides with several adverse side effects. To mitigate the adverse effects of these pesticides, investigations have focused on the development of compounds with more selectivity, and short residual life. In this research, insecticidal effects of two chitin synthesis inhibitors, hexaflumuron and lufenuron, were investigated against E. figulilella. Graded concentrations of each pesticide were prepared with distilled water. One-day-old fifth instar were sprayed by Potter's precision spray tower. Application of hexaflumuron and lufenuron on last instar larvae of E. figulilella caused not only mortality in larval stage, but also caused defects in pupal and adult stages. Larval mortality increased as concentration increased. The longevity of the fifth instars in both hexaflumuron and lufenuron treatments, in comparison with the controls, increased by more than 12 days. The longevity of adults decreased by about 10 days. Probit analysis data revealed that the sensitivity of the test insect to hexaflumuron (EC50 = 95.38 ppm) was greater than lufenuron (EC50= 379.21 ppm). PMID:23425138

Khajepour, Simin; Izadi, Hamzeh; Asari, Mohammad Javad

2012-01-01

35

Hazards and uptake of chitin synthesis inhibitors in bumblebees Bombus terrestris.  

PubMed

This research project examined the potential hazards of a major class of insect growth regulators (IGRs) to survival, reproduction and larval growth in bumblebees Bombus terrestris L. Eight chitin synthesis inhibitors (CSIs) were tested: buprofezin, cyromazine, diflubenzuron, flucycloxuron, flufenoxuron, lufenuron, novaluron and teflubenzuron. These different IGRs, which are important in the control of pest insects in greenhouses, were applied via three different routes of exposure under laboratory conditions: dermal contact, and orally via the drinking of sugar/water and via pollen. The compounds were tested at their respective maximum field recommended concentrations (MFRC) and also in dose-response assays to calculate LC(50) values. In general, none of the CSIs showed acute worker toxicity. However, there was a dramatic reduction in brood production, especially after oral treatment with pollen and sugar/water. Conspicuously, egg fertility was reduced in all treatments with diflubenzuron and teflubenzuron. In addition to egg mortality, the worker bumblebees removed larvae from the treated nest, and in most cases these individuals were dead first-second instars. Under a binocular microscope, such larvae showed an abnormally formed cuticle leading to mechanical weakness and death. In another series of experiments using (14)C-diflubenzuron and (14)C-flufenoxuron, cuticular penetration in workers was studied for a better understanding of the differences in toxicity. With (14)C-diflubenzuron, transovarial transport and accumulation in the deposited eggs supported the strong reproductive effects. Overall, the present results suggest that CSIs should be applied with caution in combination with bumblebees. The compatibility of each compound to be used in combination with B. terrestris is discussed in relation to calculated LC(50) values, routes of uptake and effects. PMID:16786494

Mommaerts, Veerle; Sterk, Guido; Smagghe, Guy

2006-08-01

36

Effects of benzoylphenylurea on chitin synthesis and orientation in the cuticle of the Drosophila larva.  

PubMed

Chitin is an essential constituent of the insect exoskeleton, the cuticle, which is an extracellular matrix (ECM) covering the animal. It is produced by the glycosyltransferase chitin synthase at the apical plasma membrane of epidermal and tracheal cells. To fulfil its role in cuticle elasticity and stiffness it associates with proteins, thereby adopting a stereotypic arrangement of helicoidally stacked sheets, which run parallel to the surface of the animal. One approach to understand the mechanisms of chitin synthesis and organisation is to dissect these processes genetically. However, since only a few genes coding for factors involved in chitin synthesis and organisation have been identified to date using the model arthropod Drosophila melanogaster insight arising from mutant analysis is rather limited. To collect new data on the role of chitin during insect cuticle differentiation, we have analysed the effects of chitin synthesis inhibitors on Drosophila embryogenesis. For this purpose, we have chosen the benzoylphenylurea diflubenzuron and lufenuron that are widely used as insect growth regulators. Our data allow mainly two important conclusions. First, correct organisation of chitin seems to directly depend on the amount of chitin synthesised. Second, chitin synthesis and organisation are cell-autonomous processes as insecticide-treated larvae display a mosaic of cuticle defects. As benzoylphenylurea are used not only as insecticides but also as anti-diabetic drugs, the study of their impact on Drosophila cuticle differentiation may be fruitful for understanding their mode of action on a cellular pathway that is seemingly conserved between vertebrates and invertebrates. PMID:18996617

Gangishetti, Umesh; Breitenbach, Sophie; Zander, Mareike; Saheb, Shaik Khaleelulla; Müller, Ursula; Schwarz, Heinz; Moussian, Bernard

2009-03-01

37

[The influence of inhibitors of neuronal and inducible NO-synthases on experimental hemorrhagic stroke.  

PubMed

Objectives. To study the effect of inhibitors of neuronal and inducible NO-synthase on the development of hemorrhagic stroke in rats Krushinsky-Molodkina (KM) without adaptation to hypoxia and with short-term adaptation to hypobaric hypoxia. Material and methods. Ninety rats were included in the study. Experiments with short-term adaptation to hypobaric hypoxia were performed on 48 rats. The inhibitor of inducible NO-synthase (aminoguanidine, "Sigma") or the inhibitor of neuronal NO-synthase (7-nitroindasol, "Sigma") were injected in dosage 2.5 mg/100g intraperitoneally. Results. Selective inhibitors of neuronal and inducible NO-synthase had a protective effect on stress injuries in KM rats. The inhibitor of neuronal NO-synthase was more effective than the inhibitor of inducible NO-synthase in the experiments without adaptation to hypoxia. Markedly greater protective effect was achieved by the simultaneous introduction of inhibitors of neuronal and inducible NO-synthase. The greatest protective effect in the development of stress damage in rats of KM was observed in short-term adaptation to hypobaric hypoxia with simultaneous introduction of both inhibitors. Conclusions. It can be assumed that an excessive amount of NO produced by neuronal and inducible NO-synthases during the acoustic exposure in KM rats leads to stress damage. Use of selective inhibitors reduce the excess NO synthesis and the development of audiogenic stress damage caused by hemorrhagic stroke. PMID:25345640

Krushinski?, A L; Kuzenkov, V S; D'iakonova, V E; Reutov, V P

2014-01-01

38

Inhibitors of Polyhydroxyalkanoate (PHA) Synthases: Synthesis, Molecular Docking, and Implications.  

PubMed

Polyhydroxyalkanoate (PHA) synthases (PhaCs) catalyze the formation of biodegradable PHAs that are considered to be ideal alternatives to non-biodegradable synthetic plastics. However, study of PhaCs has been challenging because the rate of PHA chain elongation is much faster than that of initiation. This difficulty, along with lack of a crystal structure, has become the main hurdle to understanding and engineering PhaCs for economical PHA production. Here we report the synthesis of two carbadethia CoA analogues-sT-CH2 -CoA (26?a) and sTet-CH2 -CoA (26?b)-as well as sT-aldehyde (saturated trimer aldehyde, 29), as new PhaC inhibitors. Study of these analogues with PhaECAv revealed that 26?a/b and 29 are competitive and mixed inhibitors, respectively. Both the CoA moiety and extension of PHA chain will increase binding affinity; this is consistent with our docking study. Estimation of the Kic values of 26?a and 26?b predicts that a CoA analogue incorporating an octameric hydroxybutanoate (HB) chain might facilitate the formation of a kinetically well-behaved synthase. PMID:25394180

Zhang, Wei; Chen, Chao; Cao, Ruikai; Maurmann, Leila; Li, Ping

2015-01-01

39

Glycogen synthase kinase-3 inhibitors: Rescuers of cognitive impairments  

PubMed Central

Impairment of cognitive processes is a devastating outcome of many diseases, injuries, and drugs affecting the central nervous system (CNS). Most often, very little can be done by available therapeutic interventions to improve cognitive functions. Here we review evidence that inhibition of glycogen synthase kinase-3 (GSK3) ameliorates cognitive deficits in a wide variety of animal models of CNS diseases, including Alzheimer's disease, Fragile X syndrome, Down syndrome, Parkinson's disease, spinocerebellar ataxia type 1, traumatic brain injury, and others. GSK3 inhibitors also improve cognition following impairments caused by therapeutic interventions, such as cranial irradiation for brain tumors. These findings demonstrate that GSK3 inhibitors are able to ameliorate cognitive impairments caused by a diverse array of diseases, injury, and treatments. The improvements in impaired cognition instilled by administration of GSK3 inhibitors appear to involve a variety of different mechanisms, such as supporting long-term potentiation and diminishing long-term depression, promotion of neurogenesis, reduction of inflammation, and increasing a number of neuroprotective mechanisms. The potential for GSK3 inhibitors to repair cognitive deficits associated with many conditions warrants further investigation of their potential for therapeutic interventions, particularly considering the current dearth of treatments available to reduce loss of cognitive functions. PMID:23916593

King, Margaret K.; Pardo, Marta; Cheng, Yuyan; Downey, Kimberlee; Jope, Richard S.; Beurel, Eléonore

2013-01-01

40

Chitin deacetylase product inhibition.  

PubMed

Chitin deacetylase is the only known enzyme catalyzing the hydrolysis of the acetamino linkage in the N-acetylglucosamine units of chitin and chitosan. This reaction can play an important role in enzymatic production of chitosan from chitin, or in enzymatic modification of chitosan, which has applications in medicine, pharmacy or plant protection. It was previously shown that acetic acid, a product of the deacetylation process, may act as an inhibitor of chitin deacetylase. Here we show the mechanism of inhibition of chitin deacetylase isolated from Absidia orchidis vel coerulea by acetic acid released during the deacetylation process. The process follows competitive inhibition with respect to acetic acid with an inhibition constant of K(i) = 0.286 mmol/L. These results will help to find the optimal system to carry out the enzymatic deacetylation process for industrial applications. PMID:21298809

Jaworska, Malgorzata M

2011-02-01

41

Human hematopoietic prostaglandin D synthase inhibitor complex structures.  

PubMed

In mast and Th2 cells, hematopoietic prostaglandin (PG) D synthase (H-PGDS) catalyses the isomerization of PGH(2) in the presence of glutathione (GSH) to produce the allergic and inflammatory mediator PGD(2). We determined the X-ray structures of human H-PGDS inhibitor complexes with 1-amino-4-{4-[4-chloro-6-(2-sulpho-phenylamino)-[1,3,5]triazin-2-ylmethyl]-3-sulpho-phenylamino}-9,10-dioxo-9,10-dihydro-anthracene-2-sulphonic acid (Cibacron Blue) and 1-amino-4-(4-aminosulphonyl) phenyl-anthraquinone-2-sulphonic acid (APAS) at 2.0 Å resolution. When complexed with H-PGDS, Cibacron Blue had an IC(50) value of 40 nM and APAS 2.1 ?M. The Cibacron Blue molecule was stabilized by four hydrogen bonds and ?-? stacking between the anthraquinone ring and Trp104, the ceiling of the active site H-PGDS pocket. Among the four hydrogen bonds, the Cibacron Blue terminal sulphonic group directly interacted with conserved residues Lys112 and Lys198, which recognize the PGH(2) substrate ?-chain. In contrast, the APAS anthraquinone ring was inverted to interact with Trp104, while its benzenesulphonic group penetrated the GSH-bound region at the bottom of the active site. Due to the lack of extended aromatic rings, APAS could not directly hydrogen bond with the two conserved lysine residues, thus decreasing the total number of hydrogen bond from four to one. These factors may contribute to the 50-fold difference in the IC(50) values obtained for the two inhibitors. PMID:22418579

Kado, Yuji; Aritake, Kosuke; Uodome, Nobuko; Okano, Yousuke; Okazaki, Nobuo; Matsumura, Hiroyoshi; Urade, Yoshihiro; Inoue, Tsuyoshi

2012-04-01

42

Sorting Signals That Mediate Traffic of Chitin Synthase III between the TGN/Endosomes and to the Plasma Membrane in Yeast  

PubMed Central

Traffic of the integral yeast membrane protein chitin synthase III (Chs3p) from the trans-Golgi network (TGN) to the cell surface and to and from the early endosomes (EE) requires active protein sorting decoded by a number of protein coats. Here we define overlapping signals on Chs3p responsible for sorting in both exocytic and intracellular pathways by the coats exomer and AP-1, respectively. Residues 19DEESLL24, near the N-terminal cytoplasmically-exposed domain, comprise both an exocytic di-acidic signal and an intracellular di-leucine signal. Additionally we show that the AP-3 complex is required for the intracellular retention of Chs3p. Finally, residues R374 and W391, comprise another signal responsible for an exomer-independent alternative pathway that conveys Chs3p to the cell surface. These results establish a role for active protein sorting at the trans-Golgi en route to the plasma membrane (PM) and suggest a possible mechanism to regulate protein trafficking. PMID:23056294

Wang, Chao-Wen; Schekman, Randy

2012-01-01

43

Effect of cyclooxygenase and NO synthase inhibitors on antinociceptive action of acetaminophen.  

PubMed

The influence of cyclooxygenase (COX) and NO synthase inhibitors on antinociceptive action of acetaminophen (ACETA) was studied in rats. ACETA increased the nociceptive threshold for both mechanical (Randall-Selitto test) and chemical stimuli (writhing test). In both models the existence of ceiling dose of ACETA was observed. Indomethacin (IND), an inhibitor preferentially acting on COX-1, as well as nimesulide (NIM) and celecoxib (CECOX), i.e. respectively preferential and selective inhibitors of COX-2, markedly decreased the antinociceptive activity of ACETA in Randall-Selitto test. In contrast, IND increased, whereas both NIM and CECOX did not have any effect on ACETA action in writhing test. Pretreatment with LG-nitro-L-arginine (L-NO-ARG), an unspecific inhibitor of NO synthase, 7-nitroindazole (7-NI), relatively specific inhibitor of neuronal NO synthase, and L-N6(1-iminoethyl)lysine (L-NIL), relatively selective inhibitor of inducible NO synthase, significantly increased the action of the lower doses of ACETA (50 and 100 mg/kg) in writhing test, whereas it did not modify the effects of the higher doses. Similar effect of L-NO-ARG and 7-NI was observed in Randall-Selitto test, whereas L-NIL did not influence the action of ACETA. The possible involvement of COX and NO synthase systems in antinociceptive activity of ACETA is discussed. PMID:11990080

Bujalska, M; Gumu?ka, W S

2001-01-01

44

Effects of the chitin synthesis inhibitor buprofezin on survival and development of immatures of Chrysoperla rufilabris (Neuroptera: Chrysopidae).  

PubMed

Effects of buprofezin (Applaud), a chitin synthesis inhibitor, on survival and development of eggs, three instars, and pupae of Chrysoperla rufilabris (Burmeister) were determined in the laboratory. Buprofezin at three tested concentrations (100, 500, and 1,000 mg [AI]/liter) did not affect the viability and development of eggs when the eggs were treated, or third instars and pupae when those stages were treated. Although the degree of effects by buprofezin on larvae varied with instar, buprofezin at the higher concentrations (500 and 1,000 mg [AI]/liter) reduced survival rates 17-47% and prolonged the overall development from first instars to adult emergence by 2 or 3 d when first instars were treated, indicating that the first instar is the most vulnerable stage. When second instars were treated, the survival of C. rufilabris from second instars to pupae was not significantly affected. However, the developmental time from second instar to adult emergence was longer in the treatments with the highest concentration (1,000 mg [AI]/liter) than that with the lowest concentration (100 mg [AI]/liter). The compatibility of buprofezin with natural enemies in integrated pest management programs is discussed. PMID:10826167

Liu, T X; Chen, T Y

2000-04-01

45

Behavioral and histological changes in the Formosan subterranean termite (Isoptera: Rhinotermitidae) induced by the chitin synthesis inhibitor noviflumuron.  

PubMed

This study describes the behavioral and histological changes of the molting process in Coptotermes formosanus Shiraki caused by the chitin synthesis inhibitor noviflumuron. Termites exposed to noviflumuron initiated ecdysis as untreated individuals did; however, peristalsis contractions were weak and the expansion of the dorsal breach of the exoskeleton did not occur. Treated termites could not complete their molting process and died after the initiation of the ecdysis. Histological observations showed that the process of voiding the gut protozoa during premolting was not affected by the noviflumuron treatment. However, the formation of the new cuticle was disrupted resulting in the loss of integrity of the cuticle. The alteration of the cuticle was visible in the gizzard (foregut), the thoracic pleurons, and most of the exoskeleton. Muscles were partially able to reattach to the incompletely formed new cuticle, and muscle contractions resulted in tearing off the cuticle. Because the integrity of the newly formed cuticle was compromised by the noviflumuron treatment, we concluded that termites' death was caused primarily by the loss of hemolymph as a result of the damage done by the muscle contractions on the exoskeleton during the peristalsis. As the physiological homeostasis was disrupted, termites were too weak to shed their old cuticle, ultimately resulting in termite dying during the molting process. PMID:24772556

Xing, Lin; Chouvenc, Thomas; Su, Nan-Yao

2014-04-01

46

REGULATION OF CHITIN SYNTHESIS IN THE LARVAL MIDGUT OF MANDUCA SEXTA  

Technology Transfer Automated Retrieval System (TEKTRAN)

In insects, chitin is not only synthesized by ectodermal cells that form chitinous cuticles, but also by endodermal cells of the midgut that secrete a chitinous peritrophic matrix. Using anti-chitin synthase (CHS) antibodies, we previously demonstrated that in the midgut of Manduca sexta, CHS is exp...

47

Isolation of the peptide inhibitor of H+-ATP synthase from Crithidia fasciculata and Trypanosoma cruzi.  

PubMed

An inhibitor of Crithidia fasciculata and Trypanosoma cruzi H+ -ATP synthase (ATPase) was isolated from these organims mitochondrial particles, either by (a) ammonium sulfate-cholate extraction followed by heat treatment and ethanol precipitation, or (b) gel-filtration on Sephadex G-50, followed by a similar purification procedure. Inactivation by trypsin supported the inhibitor peptide structure. Removal of the peptide inhibitor increased about three-fold the specific activity of the protozoan ATPases. The isolated peptides and a highly purified bovine heart ATPase inhibitor inhibited C. fasciculata ATPase as a function of the peptide concentration. PMID:2527503

Rilo, M C; Cataldi de Flombaum, M A; Stoppani, A O

1989-02-01

48

Chloropropionyl-CoA: a mechanism-based inhibitor of HMG-CoA synthase and fatty acid synthase  

SciTech Connect

Recent work on the mechanisms of inactivation of HMG-CoA synthase and fatty acid synthase by chloropropionyl-CoA (Cl-prop-CoA) suggests that this analog is a mechanism-based (suicide) inhibitor; the acyl group is enzymatically converted to an acrylyl derivative prior to alkylation of the target proteins. When Cl-(/sup 3/H)prop-CoA is incubated with the target enzymes, /sup 3/H/sub 2/O is produced concomitantly with enzyme inactivation; this suggests that deprotonation and chloride elimination to form an acrylyl moiety occurs. Difficulty in cleanly synthesizing acrylyl-CoA complicates direct demonstration of the intermediacy of this species. However, synthesis of a functionally equivalent reactive substrate analog, S-acrylyl-N-acetylcysteamine has been accomplished. This analog irreversibly inhibits both HMG-CoA synthase and fatty acid synthase in a site directed fashion. Concentrations required for effective inhibition (K/sub i/ values of 1.9 mM and 3.6 mM, respectively) are much higher than observed with Cl-prop-CoA. Maximal rates of inactivation (as vertical bar ..-->.. infinity) are comparable to those measured with Cl-prop-CoA, indicating that an acrylyl derivative is kinetically competent to function as an intermediate, as required if Cl-prop-CoA is a mechanism-based inhibitor. S-acrylyl-N-acetylcysteamine also inactivates HMG-CoA lyase. In this case, kinetic studies indicate that a bimolecular process is involved (k/sub 2/ = 86.7M/sup -1/min/sup -1/ at 30/sup 0/, pH 7.0).

Miziorko, H.M.; Ahmad, F.; Behnke, C.E.

1986-05-01

49

Detection of resistance to acetohydroxyacid synthase inhibitors in Amaranthus sp. using DNA polymorphisms  

Microsoft Academic Search

Resistance to acetohydroxyacid synthase inhibitors is very widespread worldwide and is generally due to various point mutations in the gene coding for the target enzyme. Rapid resistance confirmation is key for the proper management of resistance. We aimed at determining whether two DNA-based tests, PCR-RFLP and PCR amplification of specific alleles (PASA), could reliably identify common AHAS mutations in Amaranthus

Cheryl-Ann L. Corbett; François J. Tardif

2008-01-01

50

Genetic rearrangements on the Chlorovirus genome that switch between hyaluronan synthesis and chitin synthesis  

Microsoft Academic Search

Chlorella viruses or chloroviruses form polysaccharide fibers on the cell wall of host Chlorella cells after infection. Such polysaccharides are either hyaluronan synthesized by virus-encoded hyaluronan synthase (HAS) or chitin synthesized by viral chitin synthase (CHS). Some chloroviruses synthesize both hyaluronan (HA) and chitin simultaneously. To understand the relationship between “HA-synthesizing” and “chitin-synthesizing” viruses, we characterized the CVK2 genomic regions,

Ali Mohammed Mohammed Ali; Takeru Kawasaki; Takashi Yamada

2005-01-01

51

Identification, mRNA Expression, and Functional Analysis of Chitin Synthase 1 Gene and Its Two Alternative Splicing Variants in Oriental Fruit Fly, Bactrocera dorsalis  

PubMed Central

Two alternative splicing variants of chitin synthase 1 gene (BdCHS1) were cloned and characterized from the oriental fruit fly, Bactrocera dorsalis (Hendel). The cDNA of both variants (BdCHS1a and BdCHS1b) consisted of 5,552 nucleotides (nt), with an open reading frame (ORF) of 4,776 nt, encoding a protein of 1,592 amino acid residues, plus 685- and 88-nt of 5?- and 3?-noncoding regions, respectively. The alternative splicing site was located between positions 3,784-3,960 and formed a pair of mutually exclusive exons (a/b) that were same in size (177 nt), but showed only 65% identity at the nucleotide level. During B. dorsalis growth and development, BdCHS1 and BdCHS1a were both mainly expressed during the larval-pupal and pupal-adult transitions, while BdCHS1b was mainly expressed during pupal-adult metamorphosis and in the middle of the pupal stage. BdCHS1a was predominately expressed in the integument whereas BdCHS1b was mainly expressed in the trachea. The 20-hydroxyecdysone (20E) induced the expression of BdCHS1 and its variants. Injection of dsRNA of BdCHS1, BdCHS1a, and BdCHS1b into third-instar larvae significantly reduced the expression levels of the corresponding variants, generated phenotypic defects, and killed most of the treated larvae. Furthermore, silencing of BdCHS1 and BdCHS1a had a similar result in that the larva was trapped in old cuticle and died without tanning completely, while silencing of BdCHS1b has no effect on insect morphology. These results demonstrated that BdCHS1 plays an important role in the larval-pupal transition and the expression of BdCHS1 in B. dorsalis is regulated by 20E. PMID:23569438

Yang, Wen-Jia; Xu, Kang-Kang; Cong, Lin; Wang, Jin-Jun

2013-01-01

52

Structural and biological studies on bacterial nitric oxide synthase inhibitors  

PubMed Central

Nitric oxide (NO) produced by bacterial NOS functions as a cytoprotective agent against oxidative stress in Staphylococcus aureus, Bacillus anthracis, and Bacillus subtilis. The screening of several NOS-selective inhibitors uncovered two inhibitors with potential antimicrobial properties. These two compounds impede the growth of B. subtilis under oxidative stress, and crystal structures show that each compound exhibits a unique binding mode. Both compounds serve as excellent leads for the future development of antimicrobials against bacterial NOS-containing bacteria. PMID:24145412

Holden, Jeffrey K.; Li, Huiying; Jing, Qing; Kang, Soosung; Richo, Jerry; Silverman, Richard B.; Poulos, Thomas L.

2013-01-01

53

Synthesis of Bi-substrate State Mimics of Dihydropteroate Synthase as Potential Inhibitors and Molecular Probes  

PubMed Central

The increasing emergence of resistant bacteria drives us to design and develop new antimicrobial agents. Pursuant to that goal, a new targeting approach of the dihydropteroate synthase enzyme, which serves as the site of action for the sulfonamide class of antimicrobial agents, is being explored. Using structural information, a new class of transition state mimics has been designed and synthesized that have the capacity to bind to the pterin, phosphate and para-amino binding sites. The design, synthesis and evaluation of these compounds as inhibitors of Bacillus anthracis dihydropteroate synthase is described herein. Outcomes from this work have identified the first trivalent inhibitors of dihydropteroate synthase whose activity displayed slow binding inhibition. The most active compounds in this series contained an oxidized pterin ring. The binding of these inhibitors was modeled into the dihydropteroate synthase active site and demonstrated a good correlation with the observed bioassay data, as well as provided important insight for the future design of higher affinity transition state mimics. PMID:21216602

Qi, Jianjun; Virga, Kristopher G.; Das, Sourav; Zhao, Ying; Yun, Mi-Kyung; White, Stephen W.; Lee, Richard E.

2010-01-01

54

Lipophilic Bisphosphonates as Dual Farnesyl\\/Geranylgeranyl Diphosphate Synthase Inhibitors: An X-ray and NMR Investigation  

Microsoft Academic Search

Considerable effort has focused on the development of selective protein farnesyl transferase (FTase) and protein geranylgeranyl transferase (GGTase) inhibitors as cancer chemotherapeutics. Here, we report a new strategy for anticancer therapeutic agents involving inhibition of farnesyl diphosphate synthase (FPPS) and geranylgeranyl diphosphate synthase (GGPPS), the two enzymes upstream of FTase and GGTase, by lipophilic bisphosphonates. Due to dual site targeting

Yonghui Zhang; Rong Cao; Fenglin Yin; Michael P. Hudock; Rey-Ting Guo; Y Song; J No; K Bergan; A Leon; Lauren Cass; Amanda Goddard; Ting-Kai Chang; Fu-Yang Lin; Ermond Van Beek; Socrates Papapoulos; Andrew H.-J. Wang; Tadahiko Kubo; Mitsuo Ochi; Dushyant Mukkamala; Eric Oldfield

2009-01-01

55

Structures of a Potent Phenylalkyl Bisphosphonate Inhibitor Bound to Farnesyl and Geranylgeranyl Diphosphate Synthases  

PubMed Central

We report the x-ray crystallographic structures of the bisphosphonate N-[methyl(4-phenylbutyl)]-3-aminopropyl-1-hydroxy-1,1-bisphosphonate (BPH-210), a potent analog of pamidronate (Aredia), bound to farnesyl diphosphate synthase (FPPS) from Trypanosoma brucei as well as to geranylgeranyl diphosphate synthase from Saccharomyces cerevisiae. BPH-210 binds to FPPS, together with 3 Mg2+, with its long, hydrophobic phenylbutyl sidechain being located in the same binding pocket that is occupied by allylic diphosphates and other bisphosphonates. Binding is overwhelmingly entropy driven, as determined by isothermal titration calorimetry. The structure is of interest since it explains the lack of potency of longer chain analogs against FPPS, since these would be expected to have a steric clash with an aromatic ring at the distal end of the binding site. Unlike shorter chain FPPS inhibitors, such as pamidronate, BPH-210 is also found to be a potent inhibitor of human geranylgeranyl diphosphate synthase. In this case, the bisphosphonate binds only to the GGPP product inhibitory site, with only 1 (chain A) or 0 (chain B) Mg2+, and ?S is much smaller and ?H is ~6k cal more negative than in the case of FPPS binding. Overall, these results are of general interest since they show that some bisphosphonates can bind to more than one trans-prenyl synthase enzyme which, in some cases, can be expected to enhance their overall activity in vitro and in vivo. PMID:18442135

Cao, Rong; Chen, Cammy K.-M.; Guo, Rey-Ting; Wang, Andrew H.-J.; Oldfield, Eric

2008-01-01

56

Identification and characterisation of new inhibitors for the human hematopoietic prostaglandin D2 synthase.  

PubMed

Prostaglandin D(2) synthesised by the hematopoietic prostaglandin D(2) synthase has a pro-inflammatory effect in allergic asthma, regulating many hallmark characteristics of the disease. Here we describe identification of hematopoietic prostaglandin D(2) synthase inhibitors including cibacron blue, bromosulfophthalein and ethacrynic acid. Expansion around the drug-like ethacrynic acid identified a novel inhibitor, nocodazole, and a fragment representing its aromatic core. Nocodazole binding was further characterised by docking calculations in combination with conformational strain analysis. The benzyl thiophene core was predicted to be buried in the active site, binding in the putative prostaglandin binding site, and a likely hydrogen bond donor site identified. X-ray crystallographic studies supported the predicted binding mode. PMID:19939518

Weber, Jane E; Oakley, Aaron J; Christ, Angelika N; Clark, Alan G; Hayes, John D; Hall, Rhonda; Hume, David A; Board, Philip G; Smythe, Mark L; Flanagan, Jack U

2010-02-01

57

Upregulation of Endothelial Nitric Oxide Synthase by HMG CoA Reductase Inhibitors  

Microsoft Academic Search

Background—Oxidized low-density lipoprotein (ox-LDL) causes endothelial dysfunction in part by decreasing the availability of endothelial nitric oxide (NO). Although HMG CoA reductase inhibitors restore endothelial function by reducing serum cholesterol levels, it is not known whether they can also directly upregulate endothelial NO synthase (ecNOS) activity. Methods and Results—Human saphenous vein endothelial cells were treated with ox-LDL (50 mg\\/mL thiobarbituric

Ulrich Laufs; Vito La Fata; Jorge Plutzky; James K. Liao

58

Imaging Pharmacodynamics of the A-Folate Receptor-Targeted Thymidylate Synthase Inhibitor BGC 945  

Microsoft Academic Search

The assessment of tissue-specific pharmacodynamics is desirable in the development of tumor-targeted therapies. Plasma deoxyuridine (dUrd) levels, a measure of systemic thymidylate synthase (TS) inhibition, has limited applica- tion for studying the pharmacodynamics of novel TS inhibitors targeted to the high affinity A-folate receptor (FR). Here, we have evaluated the utility of (18F)fluorothy- midine positron emission tomography ((18F)FLT-PET) for imaging

Radhakrishna G. Pillai; Meg Perumal; Fraser Mitchell; Julius Leyton; Franklin I. Aibgirhio; Oksana Golovko; Ann L. Jackman; Eric O. Aboagye

59

Chitin Research Revisited  

PubMed Central

Two centuries after the discovery of chitin, it is widely accepted that this biopolymer is an important biomaterial in many aspects. Numerous studies on chitin have focused on its biomedical applications. In this review, various aspects of chitin research including sources, structure, biosynthesis, chitinolytic enzyme, chitin binding protein, genetic engineering approach to produce chitin, chitin and evolution, and a wide range of applications in bio- and nanotechnology will be dealt with. PMID:20714419

Khoushab, Feisal; Yamabhai, Montarop

2010-01-01

60

Effect of a selective thromboxane synthase inhibitor on arterial graft patency and platelet deposition in dogs  

SciTech Connect

This study examined the effect of selective thromboxane synthase inhibition and nonselective cyclooxygenase inhibition on vascular graft patency and indium 111-labeled platelet deposition in 35 mongrel dogs undergoing carotid artery replacement with 4 mm X 4 cm polytetrafluoroethylene (PTFE) (one side) and Dacron (opposite side) end-to-end grafts. Aspirin-dipyridamole therapy improved one-week graft patency, from 46% in untreated dogs to 93% in treated dogs. Thromboxane synthase inhibition (U-63557A) improved graft patency in these dogs to 81%. Both drug treatments reduced platelet deposition on Dacron and PTFE grafts by 48% to 68% compared with control dogs. Dacron grafts accumulated significantly more platelets than PTFE grafts but had comparable patency rates. Low-dose aspirin therapy had no significant effect on either graft patency or platelet deposition. All treatment groups showed a 60% to 76% reduction in serum thromboxane B2, but only thromboxane synthase inhibitor treatment increased plasma 6-keto-prostaglandin F1 alpha by 100%. Selective thromboxane synthase inhibition improved small-caliber prosthetic graft patency to the same extent as did conventional cyclooxygenase inhibition in this preliminary study.

McDaniel, M.D.; Huntsman, W.T.; Miett, T.O.; Cronenwett, J.L.

1987-08-01

61

Inhibitors of microsomal prostaglandin E2 synthase-1 enzyme as emerging anti-inflammatory candidates.  

PubMed

Cyclooxygenases (COX-1 and COX-2) catalyze the conversion of arachidonic acid (AA) into PGH2 that is further metabolized by terminal prostaglandin (PG) synthases into biologically active PGs, for example, prostaglandin E2 (PGE2), prostacyclin I2 (PGI2), thromboxane A2 (TXA2), prostaglandin D2 (PGD2), and prostaglandin F2 alpha (PGF2?). Among them, PGE2 is a widely distributed PG in the human body, and an important mediator of inflammatory processes. The successful modulation of this PG provides a beneficial strategy for the potential anti-inflammatory therapy. For instance, nonsteroidal anti-inflammatory agents (NSAIDs), both classical nonselective (cNSAIDs) and the selective COX-2 inhibitors (coxibs) attenuate the generation of PGH2 from AA that in turn reduces the synthesis of PGE2 and modifies the inflammatory conditions. However, the long-term use of these agents causes severe side effects due to the nonselective inhibition of other PGs, such as PGI2 and TXA2, etc. Microsomal prostaglandin E2 synthase-1 (mPGES-1), a downstream PG synthase, specifically catalyzes the biosynthesis of COX-2-derived PGE2 from PGH2, and describes itself as a valuable therapeutic target for the treatment of acute and chronic inflammatory disease conditions. Therefore, the small molecule inhibitors of mPGES-1 would serve as a beneficial anti-inflammatory therapy, with reduced side effects that are usually associated with the nonselective inhibition of PG biosynthesis. PMID:25019142

Bahia, Malkeet Singh; Katare, Yogesh Kumar; Silakari, Om; Vyas, Bhawna; Silakari, Pragati

2014-07-01

62

Geranyl and neryl triazole bisphosphonates as inhibitors of geranylgeranyl diphosphate synthase.  

PubMed

When inhibitors of enzymes that utilize isoprenoid pyrophosphates are based on the natural substrates, a significant challenge can be to achieve selective inhibition of a specific enzyme. One element in the design process is the stereochemistry of the isoprenoid olefins. We recently reported preparation of a series of isoprenoid triazoles as potential inhibitors of geranylgeranyl transferase II but these compounds were obtained as a mixture of olefin isomers. We now have accomplished the stereoselective synthesis of these triazoles through the use of epoxy azides for the cycloaddition reaction followed by regeneration of the desired olefin. Both geranyl and neryl derivatives have been prepared as single olefin isomers through parallel reaction sequences. The products were assayed against multiple enzymes as well as in cell culture studies and surprisingly a Z-olefin isomer was found to be a potent and selective inhibitor of geranylgeranyl diphosphate synthase. PMID:24726306

Zhou, Xiang; Ferree, Sarah D; Wills, Veronica S; Born, Ella J; Tong, Huaxiang; Wiemer, David F; Holstein, Sarah A

2014-05-01

63

ATP Synthase and the Actions of Inhibitors Utilized To Study Its Roles in Human Health, Disease, and Other Scientific Areas  

PubMed Central

Summary: ATP synthase, a double-motor enzyme, plays various roles in the cell, participating not only in ATP synthesis but in ATP hydrolysis-dependent processes and in the regulation of a proton gradient across some membrane-dependent systems. Recent studies of ATP synthase as a potential molecular target for the treatment of some human diseases have displayed promising results, and this enzyme is now emerging as an attractive molecular target for the development of new therapies for a variety of diseases. Significantly, ATP synthase, because of its complex structure, is inhibited by a number of different inhibitors and provides diverse possibilities in the development of new ATP synthase-directed agents. In this review, we classify over 250 natural and synthetic inhibitors of ATP synthase reported to date and present their inhibitory sites and their known or proposed modes of action. The rich source of ATP synthase inhibitors and their known or purported sites of action presented in this review should provide valuable insights into their applications as potential scaffolds for new therapeutics for human and animal diseases as well as for the discovery of new pesticides and herbicides to help protect the world's food supply. Finally, as ATP synthase is now known to consist of two unique nanomotors involved in making ATP from ADP and Pi, the information provided in this review may greatly assist those investigators entering the emerging field of nanotechnology. PMID:19052322

Hong, Sangjin; Pedersen, Peter L.

2008-01-01

64

Structural study and thermodynamic characterization of inhibitor binding to lumazine synthase from Bacillus anthracis  

PubMed Central

The crystal structure of lumazine synthase from Bacillus anthracis was solved by molecular replacement and refined to R cryst = 23.7% (R free = 28.4%) at a resolution of 3.5?Å. The structure reveals the icosahedral symmetry of the enzyme and specific features of the active site that are unique in comparison with previously determined orthologues. The application of isothermal titration calorimetry in combination with enzyme kinetics showed that three designed pyrimidine derivatives bind to lumazine synthase with micromolar dissociation constants and competitively inhibit the catalytic reaction. Structure-based modelling suggested the binding modes of the inhibitors in the active site and allowed an estimation of the possible contacts formed upon binding. The results provide a structural framework for the design of antibiotics active against B. anthracis. PMID:20823551

Morgunova, Ekaterina; Illarionov, Boris; Saller, Sabine; Popov, Aleksander; Sambaiah, Thota; Bacher, Adelbert; Cushman, Mark; Fischer, Markus; Ladenstein, Rudolf

2010-01-01

65

The Natural Product Noformycin Is an Inhibitor of Inducible-Nitric Oxide Synthase  

Microsoft Academic Search

Inducible-Nitric oxide synthase (iNOS, EC 1.14.13.39) catalyzes the formation of nitric oxide (NO) andL-citrulline fromL-Arg, NADPH and dioxygen. The natural product, (?)-noformycin was found to be a potent, competitive inhibitor of recombinant human iNOS with respect toL-Arg with aKi= 1.3 ± 0.3 ?M. The reversible binding of noformycin caused a high spin type I spectral perturbation of the iNOS heme

Barbara Gordon Green; Renee Chabin; Stephan K. Grant

1996-01-01

66

Anmindenols A and B, inducible nitric oxide synthase inhibitors from a marine-derived Streptomyces sp.  

PubMed

Anmindenols A (1) and B (2), inhibitors of inducible nitric oxide synthase (iNOS), were isolated from a marine-derived bacterium Streptomyces sp. Their chemical structures were elucidated by interpreting various spectroscopic data, including IR, MS, and NMR. Anmindenols A and B are sesquiterpenoids possessing an indene moiety with five- and six-membered rings derived from isoprenyl units. The absolute configuration of C-4 in anmindenol B was determined by electronic circular dichroism (ECD) of a dimolybdenum complex. Anmindenols A (1) and B (2) inhibited nitric oxide production in stimulated RAW 264.7 macrophage cells with IC50 values of 23 and 19 ?M, respectively. PMID:24878306

Lee, Jihye; Kim, Hiyoung; Lee, Tae Gu; Yang, Inho; Won, Dong Hwan; Choi, Hyukjae; Nam, Sang-Jip; Kang, Heonjoong

2014-06-27

67

Aldosterone synthase inhibitors as promising treatments for mineralocorticoid dependent cardiovascular and renal diseases.  

PubMed

Besides the well-known roles of aldosterone as a mineralocorticoid in regulating homeostasis of electrolytes and volume, recent studies revealed that it is also a potent proinflammation factor inducing reactive oxygen species and up-regulating a panel of fibrosis related genes. Under pathological circumstances, excessive aldosterone is involved in a lot of chronic diseases, including hypertension, cardiac fibrosis, congestive heart failure, ventricular remodeling, and diabetic nephropathy. Therefore, the inhibition of aldosterone synthase (CYP11B2), which is the pivotal enzyme in aldosterone biosynthesis, was proposed as a superior approach. Expected pharmacodynamic effects have been demonstrated in both animal models and clinical trials after the application of CYP11B2 inhibitors. The importance of selectivity over other steroidogenic CYP enzymes, in particular 11?-hydroxylase (CYP11B1), was also revealed. Recently, much more selective CYP11B2 inhibitors have been reported, which could be promising drug candidates for the treatment of aldosterone related diseases. PMID:24422519

Hu, Qingzhong; Yin, Lina; Hartmann, Rolf W

2014-06-26

68

A human fatty acid synthase inhibitor binds ?-ketoacyl reductase in the keto-substrate site.  

PubMed

Human fatty acid synthase (hFAS) is a complex, multifunctional enzyme that is solely responsible for the de novo synthesis of long chain fatty acids. hFAS is highly expressed in a number of cancers, with low expression observed in most normal tissues. Although normal tissues tend to obtain fatty acids from the diet, tumor tissues rely on de novo fatty acid synthesis, making hFAS an attractive metabolic target for the treatment of cancer. We describe here the identification of GSK2194069, a potent and specific inhibitor of the ?-ketoacyl reductase (KR) activity of hFAS; the characterization of its enzymatic and cellular mechanism of action; and its inhibition of human tumor cell growth. We also present the design of a new protein construct suitable for crystallography, which resulted in what is to our knowledge the first co-crystal structure of the human KR domain and includes a bound inhibitor. PMID:25086508

Hardwicke, Mary Ann; Rendina, Alan R; Williams, Shawn P; Moore, Michael L; Wang, Liping; Krueger, Julie A; Plant, Ramona N; Totoritis, Rachel D; Zhang, Guofeng; Briand, Jacques; Burkhart, William A; Brown, Kristin K; Parrish, Cynthia A

2014-09-01

69

Tritritionikkomycin Z, (uracil-5- sup 3 H,pyridyl-2,4- sup 3 H sub 2 ): Radiolabeling of a potent inhibitor of fungal and insect chitin synthetase  

SciTech Connect

Nikkomycin Z (NZ) (a potent fungicide, insecticide, miticide, and inhibitor of fungal and insect chitin synthetase) was converted to a mixture of specific mono-, di-, and tribromo derivatives (BrNZ, Br{sub 2}NZ, and Br{sub 3}NZ, respectively) on reaction with N-bromosuccinimide in N,N-dimethylformamide. Substitution by bromine occurred first at the 2-position of the 3-hydroxypyridyl moiety, second at the 5-position of the uracil moiety, and finally at the 4-position of the 3-hydroxypyridyl moiety as observed both for NZ and for mixtures of uridine and 3-hydroxy-6-methylpyridine as model compounds representative of the moieties of NZ. Following fractionation of the various bromonikkomycin derivatives by HPLC, their structures were assigned by NMR, MS, and UV analyses. Catalytic reductive debromination of Br{sub 3}NZ with tritium gas over palladium on carbon gave (uracil-5-{sup 3}H,pyridyl-2,4-{sup 3}H{sub 2})NZ. This material has sufficiently high specific activity ({approximately}60 Ci/mmol) and suitable positions of labeling to study its uptake, distribution, metabolism, and possible target site interactions in fungal and insect systems.

Ando, Tetsu; Tecle, B.; Toia, R.F.; Casida, J.E. (Univ. of California, Berkeley (USA))

1990-08-01

70

Glycogen Synthase Kinase-3 Inhibitors as Potent Therapeutic Agents for the Treatment of Parkinson Disease.  

PubMed Central

Parkinson's disease (PD) is a devastating neurodegenerative disorder characterized by degeneration of the nigrostriatal dopaminergic pathway. Because the current therapies only lead to temporary, limited improvement and have severe side effects, new approaches to treat PD need to be developed. To discover new targets for potential therapeutic intervention, a chemical genetic approach involving the use of small molecules as pharmacological tools has been implemented. First, a screening of an in-house chemical library on a well-established cellular model of PD was done followed by a detailed pharmacological analysis of the hits. Here, we report the results found for the small heterocyclic derivative called SC001, which after different enzymatic assays was revealed to be a new glycogen synthase kinase-3 (GSK-3) inhibitor with IC50 = 3.38 ± 0.08 ?M. To confirm that GSK-3 could be a good target for PD, the evaluation of a set of structurally diverse GSK-3 inhibitors as neuroprotective agents for PD was performed. Results show that inhibitors of GSK-3 have neuroprotective effects in vitro representing a new pharmacological option for the disease-modifying treatment of PD. Furthermore, we show that SC001 is able to cross the blood–brain barrier, protects dopaminergic neurons, and reduces microglia activation in in vivo models of Parkinson disease, being a good candidate for further drug development. PMID:23421686

2012-01-01

71

Chitin in pogonophore tubes  

Microsoft Academic Search

The chitin from pogonophore tubes is shown to be essentially the same as that from the pen of the squid Loligo and is classified as \\/I-chitin (as are the known chitins in annelids). However, certain important differences in detail are seen when comparing X-ray diffraction photographs and infra-red absorption spectra of the chitin from the two sources. These matters are

J. Blackwell; K. D. Parker; K. M. Rudall

1965-01-01

72

The glucosylceramide synthase inhibitor PDMP sensitizes pancreatic cancer cells to MEK/ERK inhibitor AZD-6244.  

PubMed

Here we show that d,l-Threo-1-phenyl-2-decanoylamino-3-morpholino-1-propanol (PDMP), a glycosphingolipid biosynthesis inhibitor, increases the sensitivity of pancreatic cancer cells to the novel MEK-ERK inhibitor AZD-6244. AZD-6244 and PDMP co-administration induced massive pancreatic cancer cell death and apoptosis, more potently than either drug alone. We discovered that AZD-6244 induced ceramide production in pancreatic cancer cells, yet the excess ceramide was metabolically removed in the long-term (24-48h). PDMP facilitated AZD-6244-induced ceramide production, and ceramide level remained elevated up to 48h. Meanwhile, exogenously-added cell-permeable short chain ceramide (C2) similarly sensitized AZD-6244's activity, the two caused substantial pancreatic cancer cell death and apoptosis. At the molecular level, PDMP and AZD-6244 co-treatment inactivated ERK1/2 and AKT-mTOR signalings simultaneously in pancreatic cancer cells, while either agent alone only affected one signaling. In summary, PDMP significantly increased the sensitivity of AZD-6244 in pancreatic cancer cells. This appears to involve a sustained ceramide production as well as concurrent block of ERK and AKT-mTOR signalings. PMID:25498501

Wang, Ting; Wei, Jue; Wang, Na; Ma, Jia-Li; Hui, Ping-Ping

2015-01-16

73

Synthesis and biological evaluation of potential threonine synthase inhibitors: Rhizocticin A and Plumbemycin A.  

PubMed

Rhizocticins and Plumbemycins are natural phosphonate antibiotics produced by the bacterial strains Bacillus subtilis ATCC 6633 and Streptomyces plumbeus, respectively. Up to now, these potential threonine synthase inhibitors have only been synthesized under enzymatic catalysis. Here we report the chemical stereoselective synthesis of the non-proteinogenic (S,Z)-2-amino-5-phosphonopent-3-enoic acid [(S,Z)-APPA] and its use for the synthesis of Rhizocticin A and Plumbemycin A. In this work, (S,Z)-APPA was synthesized via the Still-Gennari olefination starting from Garner's aldehyde. The Michaelis-Arbuzov reaction was used to form the phosphorus-carbon bond. Oligopeptides were prepared using liquid phase peptide synthesis (LPPS) and were tested against selected bacteria and fungi. PMID:23891162

Gahungu, Mathias; Arguelles-Arias, Anthony; Fickers, Patrick; Zervosen, Astrid; Joris, Bernard; Damblon, Christian; Luxen, André

2013-09-01

74

Protective effect of inducible nitric oxide synthase inhibitor on pancreas transplantation in rats  

PubMed Central

AIM: To investigate the effect of inducible nitric oxide synthase inhibitor, aminoguanidine, on pancreas transplantation in rats. METHODS: A model of pancreas transplantation was established in rats. Streptozotocin-induced diabetic male Wistar rats were randomly assigned to sham-operation control group (n = 6), transplant control group (n = 6), and aminoguanidine (AG) treatment group (n = 18). In the AG group, aminoguanidine was added to intravascular infusion as the onset of reperfusion at the dose of 60 mg/kg, 80 mg/kg, 100 mg/kg body weight, respectively. Serum nitric oxide (NO) level, blood sugar and amylase activity were detected. Nitric oxide synthase (NOS) test kit was used to detect the pancreas cNOS and inducible NOS (iNOS) activity. Pancreas sections stained with HE and immunohistochemistry were evaluated under a light microscope. RESULTS: As compared with the transplant control group, the serum NO level and amylase activity decreased obviously and the evidence for pancreas injury was much less in the AG group. The AG (80 mg/kg body weight) group showed the most significant difference in NO and amylase (NO: 66.0 ± 16.6 vs 192.3 ± 60.0, P < 0.01 and amylase: 1426 ± 177 vs 4477 ± 630, P < 0.01).The expression and activity of tissue iNOS, and blood sugar in the AG (80 mg/kg body weight) group were much lower than those in the transplant control group (iNOS: 2.01 ± 0.23 vs 26.59 ± 5.78, P < 0.01 and blood sugar: 14.2 ± 0.9 vs 16.8 ± 1.1, P < 0.01). CONCLUSION: Selective iNOS inhibitor, aminoguanidine as a free radical, has a protective effect on pancreas transplantation in rats by inhibiting NO and reducing its toxicity. PMID:18023101

Li, Bai-Feng; Liu, Yong-Feng; Cheng, Ying; Zhang, Ke-Zhong; Li, Tie-Min; Zhao, Ning

2007-01-01

75

Area Wide Field Study on Effect of Three Chitin Synthesis Inhibitor Baits on Populations of Coptotermes formosanus Shiraki and Reticulitermes flavipes (Kollar) (Isoptera: Rhinotermitidae)  

Technology Transfer Automated Retrieval System (TEKTRAN)

Periodic sampling of 43 independent monitors, initially active with Formosan subterranean termite, Coptotermes formosanus Shiraki, or the eastern subterranean termite, Reticulitermes flavipes (Kollar) was conducted to evaluate the effects of cellulose baits containing one of three chitin synthesis i...

76

Effect of cyclooxygenase and NO synthase inhibitors administered centrally on antinociceptive action of acetaminophen (Part II).  

PubMed

As it has been demonstrated in a previous study, both cyclooxygenases (COXs) and nitric oxide synthases (NOSs) participate in the mechanism of acetaminophen (ACETA) action. Results obtained in this study indicate that intrathecal (it) or intracerebroventricular (icv) pretreatment with LG-nitro-L-arginine (L-NO-Arg), a non-selective inhibitor of NOS activity, as well as with 7-nitroindazole (7-NI), a selective nNOS inhibitor, potentiated the antinociceptive activity of subceiling doses of ACETA, but were without effect on the action of supramaximal doses in Randall-Selitto test. Similar effect of L-NO-Arg and 7-NI it was observed in writhing test, whereas L-NO-Arg icv or L-NIL it did not influence the action of ACETA in this model. Indomethacin (IND), an inhibitor preferentially acting on COX-1, as well as nimesulide (NIM) and celecoxib (CECOX), i.e. preferential and selective inhibitor of COX-2, respectively, administered icv almost completely blocked the antinociceptive effect of ACETA in Randall-Selitto method. On the other hand, pretreatment with NSAIDs it initially increased and then attenuated the ACETA antinociception. Yohimbine (YOH), an alpha2-adrenergic receptor antagonist, did not modify the antinociceptive action of ACETA administered alone. However, YOH decreased the nociceptive threshold increased by simultaneously administered IND and ACETA, NIM and ACETA, as well as CECOX and ACETA in Randall-Selitto model. In contrast to the peripheral (sc) application, IND administered centrally (icv or it) did not modify the ACETA antinociception in writhing test. Neither NIM nor CECOX administered sc, it or icv changed the ACETA antinociception in this model. Possible mechanisms and sites of antinociceptive effects of ACETA are discussed. PMID:14730095

Bujalska, Magdalena

2003-01-01

77

Effects of Sublethal Concentrations of the Chitin Synthesis Inhibitor, Hexaflumuron, on the Development and Hemolymph Physiology of the Cutworm, Spodoptera litura  

PubMed Central

The effects of sublethal concentrations 0.1, 0.5, and 1.2 µg mL-1of the chitin synthesis inhibitor, hexaflumuron, on larval growth and development, the count and proportion of hemocytes, and carbohydrate content (trehalose and glyceride) in hemolymph were investigated in the cutworm, Spodoptera litura (Fabricious) (Lepidoptera: Noctuidae). When 3rdinstar larvae were subjected to the sublethal concentrations, there were dose-dependent effects on larval weight and length of each instar larvae, percent pupation and the duration of development. Most of the larvae died during the molting process at all concentrations. Few individuals from 0.5 and 1.2 µg mL -1concentrations could develop to the 6thinstar, while the pupae emerging from the 0.1 µg mL -1concentrations did not exceed 16% of the number of the initial larvae. In 5thinstar S. litura, the total number of hemocytes was significantly increased at 24 hours post—treatment, whereas the proliferation of hemocytes was inhibited, plasmatocyte pseudopodia contracted, and granulocyte expanded at 96 hours post—treatment. The increases of plasmatocyte count and the decreases of granulocyte count were dose—dependent. The longer treatment time of the sublethal concentrations increased the content of total carbohydrate and trehalose in hematoplasma, and was dose—dependent in hemocytes. The content of glyceride in hemolymph was significantly higher at 24 hours post—treatment, but gradually returned to normal levels at 96 hours post—treatment as compared with the control. The results suggested that sublethal concentrations of hexaflumuron reduced S. litura larval survival and interfered with hemolymph physiological balances. PMID:22958164

Zhu, Qiqi; He, Yuan; Yao, Jing; Liu, Yinzhao; Tao, Liming; Huang, Qingchun

2012-01-01

78

Polymorphic human prostaglandin H synthase-2 proteins and their interactions with cyclooxygenase substrates and inhibitors.  

PubMed

The cyclooxygenase (COX) activity of prostaglandin H synthase-2 (PGHS-2) is implicated in colorectal cancer and is targeted by nonsteroidal anti-inflammatory drugs (NSAIDs) and dietary n-3 fatty acids. We used purified, recombinant proteins to evaluate the functional impacts of the R228H, E488G, V511A and G587R PGHS-2 polymorphisms on COX activity, fatty acid selectivity and NSAID actions. Compared to wild-type PGHS-2, COX activity with arachidonate was ?20% lower in 488G and ?20% higher in 511A. All variants showed time-dependent inhibition by the COX-2-specific inhibitor (coxib) nimesulide, but 488G and 511A had 30-60% higher residual COX activity; 511A also showed up to 70% higher residual activity with other time-dependent inhibitors. In addition, 488G and 511A differed significantly from wild type in Vmax values with the two fatty acids: 488G showed ?20% less and 511A showed ?20% more discrimination against eicosapentaenoic acid. The Vmax value for eicosapentaenoate was not affected in 228H or 587R, nor were the Km values or the COX activation efficiency (with arachidonate) significantly altered in any variant. Thus, the E488G and V511A PGHS-2 polymorphisms may predict who will most likely benefit from interventions with some NSAIDs or n-3 fatty acids. PMID:20548327

Liu, W; Poole, E M; Ulrich, C M; Kulmacz, R J

2011-10-01

79

Differential suppression of glial nitric oxide synthase induction by structurally related tyrosine kinase inhibitors.  

PubMed

Incubation of C6 astrocytoma cells with bacterial endotoxin (lipopolysaccharide; LPS) plus interferon-gamma (IFN-gamma), or with a combination of cytokines (TNF-alpha, IL1-beta, and IFN-gamma) leads to high levels of inducible nitric oxide synthase (iNOS) expression. Previous results demonstrated a requirement for tyrosine kinase (TK) activities for iNOS induction. In the present study, a set of structurally related TK inhibitors, the tyrphostins (TYRs), were used to characterize possible differences between LPS and cytokine iNOS induction. All TYRs tested suppressed both types of induction. However, dose-response curves revealed significant differences in the IC50 values obtained for some TYRs (T25 and T56), and significant differences in the IC50 potency rank order when comparing inhibition of LPS versus cytokine-dependent iNOS induction. These results are consistent with differential TK utilization by the LPS versus cytokine pathways of iNOS induction, and establish a basis for developing further selective inhibitors of iNOS expression. PMID:9064610

Galea, E; Reddi, J; Feinstein, D L

1995-11-24

80

A high-throughput screening assay for identification of inhibitors of the A1AO-ATP synthase of the rumen methanogen Methanobrevibacter ruminantium M1.  

PubMed

We report the development of a high-throughput screening platform to identify inhibitors of the membrane-bound A1Ao-ATP synthase from the rumen methanogen Methanobrevibacter ruminantium M1. Inhibitors identified in the screen were tested against growing cultures of M. ruminantium, validating the approach to identify new inhibitors of methanogens. PMID:25575416

Aung, Htin Lin; Dey, Debjit; Janssen, Peter H; Ronimus, Ron S; Cook, Gregory M

2015-03-01

81

In vivo study of radioprotective effect of NO-synthase inhibitors and acetyl-L-carnitine.  

PubMed

This study investigated the protective effect of two nitric oxide synthase inhibitors N(omega)-nitro-L-arginine methyl ester (L-NAME, 100 mg/kg i.p.) and aminoguanidine (AG, 400 mg/kg i.p.), and an antioxidant acetyl-L-carnitine (ALC, 250 mg/kg i.p., once daily for five days) against radiation-induced damage in Wistar rats. Blood samples were collected 6 h after whole-body irradiation with 8 Gy. Plasma concentrations of nitrite+nitrate (NO(x)) and malondialdehyde (MDA) were measured by high-performance liquid chromatography. A single injection of L-NAME one hour before exposure effectively prevented the radiation-induced elevation of plasma NO(x) and it reduced 2.6-fold the risk for death during the subsequent 30-day period. Pretreatment with ALC prevented the radiation-induced increase in plasma MDA and it had similar effect on mortality as L-NAME did. Presumably due to its short half-life, the partially iNOS-selective inhibitor and antioxidant AG given in a single dose before exposure did not attenuate MDA and NO(x) and it failed to significantly improve the 30-day survival. In conclusion, pretreatment with both the nonspecific NOS inhibitor L-NAME and the antioxidant ALC markedly reduce mortality to radiation sickness in rats. The radioprotective effect may be directly related to effective attenuation of the radiation-induced elevation of NO production by L-NAME and of oxidative stress by ALC. PMID:23869893

Babicová, A; Havlínová, Z; Hroch, M; Rezá?ová, M; Pejchal, J; Vávrová, J; Chládek, J

2013-12-20

82

Biochemical characterization and evaluation of potent inhibitors of the Pseudomonas aeruginosa PA01 acetohydroxyacid synthase.  

PubMed

Microbes and plants synthesize essential branched-chain amino acids (BCAAs) such as valine, leucine, and isoleucine via a common biosynthetic pathway in which the first reaction is catalyzed by acetohydroxyacid synthase (AHAS, EC 4.1.3.18). Recently, AHAS was identified as a potential anti bacterial target. To help find an effective inhibitor that could act as an antibacterial compound, we cloned and characterized the catalytic subunit (CSU) of Pseudomonas aeruginosa AHAS, and found four potent inhibitors through chemical library screening. The ilvI gene of P. aeruginosa encodes a 65-kDa AHAS protein, consistent with the size of the purified enzyme on SDS-PAGE. Enzyme kinetics showed that the enzyme has a Km of 14.2 mM and a specific activity of 0.12 U/mg. Enzyme activity was optimum at a temperature of 37 °C and a pH of 7.5. The Kd for thiamine diphosphate (ThDP) was 89.92 ± 17.9 ?M, as determined by fluorescence quenching. The cofactor activation constants (Ks) for ThDP and (Kc) for Mg(2+) were 0.6 ± 0.1 and 560.8 ± 7.4 ?M, respectively. Further, we determined that AVS2087, AVS2093, AVS2236, and AVS2387 compounds are potent inhibitors of the catalytic subunit of P. aeruginosa AHAS. These compounds inhibit nearly 100% of AHAS activity, with IC50 values of 1.19 ?M, 5.0 nM, 25 nM, and 13 nM, respectively. Compound AVS2093 showed growth inhibition with a minimal inhibitory concentration (MIC) of 742.9 ?g/ml against P. aeruginosa strain ATCC 9027. Furthermore, these findings were supported by molecular docking studies with the AVS compounds against P. aeruginosa AHAS in which AVS2093 showed minimum binding energy (-7.8 kJ/mol) by interacting with the receptor through a single hydrogen bond of 2.873 ?. Correlation of AVS2093 activity with P. aeruginosa AHAS cell growth inhibition suggested that AHAS might serve as a target protein for the development of novel antibacterial therapeutics. Results of the current study provide an impetus to further evaluate the potency of these inhibitors against pathogenic P. aeruginosa strains in vivo and to design more potent antibacterial agents based on these AVS inhibitors. PMID:23523771

Cho, June-Haeng; Lee, Mi-Young; Baig, Irshad Ahmed; Ha, Na-Reum; Kim, Joungmok; Yoon, Moon-Young

2013-07-01

83

Nitric Oxide Synthase Inhibitor Improves De Novo and Long-Term l-DOPA-Induced Dyskinesia in Hemiparkinsonian Rats  

PubMed Central

Inhibitors of neuronal and endothelial nitric oxide synthase decrease l-3,4-dihidroxifenilalanine (l-DOPA)-induced dyskinesias in rodents. The mechanism of nitric oxide inhibitor action is unknown. The aims of the present study were to investigate the decrease of l-DOPA-induced abnormal involuntary movements (AIMs) in 6-hydroxydopamine (6-OHDA)-lesioned rats by nitric oxide inhibitors following either acute or chronic treatment. The primary findings of this study were that NG-nitro-l-Arginine, an inhibitor of endothelial and neuronal nitric oxide synthase, attenuated AIMs induced by chronic and acute l-DOPA. In contrast, rotational behavior was attenuated only after chronic l-DOPA. The 6-OHDA lesion and the l-DOPA treatment induced a bilateral increase (1.5 times) in the neuronal nitric oxide synthase (nNOS) protein and nNOS mRNA in the striatum and in the frontal cortex. There was a parallel increase, bilaterally, of the FosB/?FosB, primarily in the ipsilateral striatum. The exception was in the contralateral striatum and the ipsilateral frontal cortex, where chronic l-DOPA treatment induced an increase of approximately 10 times the nNOS mRNA. Our results provided further evidence of an anti-dyskinetic effect of NOS inhibitor. The effect appeared under l-DOPA acute and chronic treatment. The l-DOPA treatment also revealed an over-expression of the neuronal NOS in the frontal cortex and striatum. Our results corroborated findings that l-DOPA-induced rotation differs between acute and chronic treatment. The effect of the NOS inhibitor conceivably relied on the l-DOPA structural modifications in the Parkinsonian brain. Taken together, these data provided a rationale for further evaluation of NOS inhibitors in the treatment of l-DOPA-induced dyskinesia. PMID:21713068

Padovan-Neto, Fernando Eduardo; Echeverry, Marcela Bermúdez; Chiavegatto, Silvana; Del-Bel, Elaine

2011-01-01

84

Lipid-lowering properties of TAK-475, a squalene synthase inhibitor, in vivo and in vitro.  

PubMed

1. Squalene synthase is the enzyme that converts farnesyl pyrophosphate to squalene in the cholesterol biosynthesis pathway. We examined the lipid-lowering properties of 1-[[(3R,5S)-1-(3-acetoxy-2,2-dimethylpropyl)-7-chloro-5-(2,3-dimethoxyphenyl)-2-oxo-1,2,3,5-tetrahydro-4,1-benzoxazepin-3-yl]acetyl]piperidine-4-acetic acid (TAK-475), a novel squalene synthase inhibitor. 2. TAK-475 inhibited hepatic cholesterol biosynthesis in rats (ED(50), 2.9 mg kg(-1)) and showed lipid-lowering effects in beagle dogs, marmosets, cynomolgus monkeys and Wistar fatty rats. 3. In marmosets, TAK-475 (30, 100 mg kg(-1), p.o., for 4 days) lowered both plasma non-high-density lipoprotein (HDL) cholesterol and triglyceride, but did not affect plasma HDL cholesterol. On the other hand, atorvastatin (10, 30 mg kg(-1), p.o., for 4 days) lowered the levels of all these lipids. A correlation between decrease in triglyceride and increase in HDL cholesterol was observed, and TAK-475 increased HDL cholesterol with a smaller decrease in triglyceride than did atorvastatin. 4. TAK-475 (60 mg kg(-1), p.o., for 15 days) suppressed the rate of triglyceride secretion from the liver in hypertriglyceridemic Wistar fatty rats, which show an enhanced triglyceride secretion rate from the liver compared with their lean littermates. 5. In HepG2 cells, TAK-475 and its pharmacologically active metabolite, T-91485, increased the binding of (125)I-low-density lipoprotein (LDL) to LDL receptors. 6. These results suggest that TAK-475 has clear hypolipidemic effects in animals via inhibition of hepatic triglyceride secretion and upregulation of LDL receptors, and that TAK-475 might increase HDL cholesterol by decreasing triglyceride. Thus, TAK-475 is expected to be useful for the treatment of dyslipidemia. PMID:12839864

Nishimoto, Tomoyuki; Amano, Yuichiro; Tozawa, Ryuichi; Ishikawa, Eiichiro; Imura, Yoshimi; Yukimasa, Hidefumi; Sugiyama, Yasuo

2003-07-01

85

Non-antibiotic quorum sensing inhibitors acting against N-acyl homoserine lactone synthase as druggable target  

PubMed Central

N-acylhomoserine lactone (AHL)-based quorum sensing (QS) is important for the regulation of proteobacterial virulence determinants. Thus, the inhibition of AHL synthases offers non-antibiotics-based therapeutic potentials against QS-mediated bacterial infections. In this work, functional AHL synthases of Pseudomonas aeruginosa LasI and RhlI were heterologously expressed in an AHL-negative Escherichia coli followed by assessments on their AHLs production using AHL biosensors and high resolution liquid chromatography–mass spectrometry (LCMS). These AHL-producing E. coli served as tools for screening AHL synthase inhibitors. Based on a campaign of screening synthetic molecules and natural products using our approach, three strongest inhibitors namely are salicylic acid, tannic acid and trans-cinnamaldehyde have been identified. LCMS analysis further confirmed tannic acid and trans-cinnemaldehyde efficiently inhibited AHL production by RhlI. We further demonstrated the application of trans-cinnemaldehyde inhibiting Rhl QS system regulated pyocyanin production in P. aeruginosa up to 42.06%. Molecular docking analysis suggested that trans-cinnemaldehyde binds to the LasI and EsaI with known structures mainly interacting with their substrate binding sites. Our data suggested a new class of QS-inhibiting agents from natural products targeting AHL synthase and provided a potential approach for facilitating the discovery of anti-QS signal synthesis as basis of novel anti-infective approach. PMID:25430794

Chang, Chien-Yi; Krishnan, Thiba; Wang, Hao; Chen, Ye; Yin, Wai-Fong; Chong, Yee-Meng; Tan, Li Ying; Chong, Teik Min; Chan, Kok-Gan

2014-01-01

86

Lipophilic 1,1-bisphosphonates are potent squalene synthase inhibitors and orally active cholesterol lowering agents in vivo.  

PubMed

Squalene synthase catalyzes the reductive dimerization of two molecules of farnesyl diphosphate to form squalene at the final branchpoint of the cholesterol biosynthetic pathway. We report herein that isoprenyl 1,1-bisphosphonates and related analogs are potent inhibitors of rat microsomal squalene synthase (I50 = 0.7-32 nM). In addition, members of this family are potent inhibitors of cholesterol biosynthesis in rats on intravenous and oral dosing, as well as cholesterol lowering agents in rats and hamsters. Significant inhibition of cholesterol biosynthesis in rats by lovastatin occurs with a concomitant inhibition of dolichol and coenzyme-Q9 synthesis. In contrast, bisphosphonate 4 has no effect on dolichol and coenzyme-Q9 biosynthesis in rats under conditions where cholesterol biosynthesis is > 90% inhibited. PMID:8227045

Ciosek, C P; Magnin, D R; Harrity, T W; Logan, J V; Dickson, J K; Gordon, E M; Hamilton, K A; Jolibois, K G; Kunselman, L K; Lawrence, R M

1993-11-25

87

Imaging Pharmacodynamics of the alpha Folate Targeted Thymidylate Synthase Inhibitor BGC 945  

PubMed Central

The assessment of tissue-specific pharmacodynamics is desirable in the development of tumour-targeted therapies. Plasma deoxyuridine (dUrd) levels - a measure of systemic thymidylate synthase (TS) inhibition - has limited application for studying the pharmacodynamics of novel TS inhibitors targeted to the high affinity alpha-folate receptor (FR). Here we have evaluated the utility of [18F]fluorothymidine positron emission tomography ([18F]FLT-PET) for imaging the tissue pharmacodynamics of BGC 945, a FR-targeted antifolate TS inhibitor; the non-targeted antifolate BGC 9331 was used for comparison. TS inhibition by both drugs induced a concentration-dependent increase in [3H]thymidine uptake in FR-positive human epidermoid KB cells. Membrane associated equilibrative nucleoside transporter type 1 levels increased from 55720±6101 to 118700±5193 and 130800±10800 per cell at 100 ?g/ml of BGC 9331 and BGC 945, respectively, suggesting this as a potential mechanism of increased nucleoside uptake. In keeping with these in vitro findings, tumour [18F]FLT accumulation in KB xenografts increased by ?2-fold after drug treatment with maximal levels at 1-4 and 4-24 h following BGC 9331 and BGC 945 treatment, respectively. Of interest to FR-targeting, BGC 9331 but not BGC 945 induced accumulation of [18F]FLT uptake in intestine, a proliferative and TS-responsive tissue. For both drugs, quantitative changes in tumour [18F]FLT uptake were associated with increased tumour dUrd levels. In conclusion, we have validated the utility of [18F]FLT-PET to image TS inhibition induced by antifolates and demonstrated the tumour-specific activity of BGC 945. This imaging biomarker readout will be useful in the early clinical development of BGC 945. PMID:18483267

Pillai, Radhakrishna G.; Forster, Martin; Perumal, Meg; Mitchell, Fraser; Leyton, Julius; Aibgirhio, Franklin I.; Golovko, Oksana; Jackman, Ann L.; Aboagye, Eric O.

2008-01-01

88

Alterations on the growth and ultrastructure of Leishmania chagasi induced by squalene synthase inhibitors.  

PubMed

Leishmaniasis is an important disease in widely dispersed regions of the world. In South America, visceral leishmaniasis (VL) is mainly caused by Leishmania chagasi. The morbidity associated with the infection is high, and death may occur in some untreated patients. Treatment has been based upon pentavalent antimonial drugs for more than half a century and problems, including development of resistance to antimonials and lack of efficacy against VL/HIV co-infections, have emphasized the need for new drugs. Squalene synthase (SQS) is an essential enzyme for the biosynthesis of protozoal sterol molecules. In this work, nineteen synthetic quinuclidines, potentially inhibitors of SQS, were tested against promastigote forms of L. chagasi and the IC50 values of the compounds were determined. The most active compounds had IC50 values of around 30 nM and induced complete growth arrest and cell lysis at sub-micromolar concentrations. We analyzed the morphological structure of the parasites treated with these compounds by transmission electron microscopy of thin sections. Treated parasites showed significant ultrastructural changes, which varied from discrete alterations to total destruction of the cells, depending on the drug concentration and the time of incubation. One important change observed was a typical swelling of the unique and highly branched mitochondrion, where the inner membrane lost its organization. There was an increase in the number of autophagosomal structures. Changes in the organization of the nuclear chromatin and alterations in the flagellar pocket and flagellar membrane were also observed. PMID:17367936

Granthon, Ana Claudia; Braga, Marina V; Rodrigues, Juliany C F; Cammerer, Simon; Lorente, Silvia Orenes; Gilbert, Ian H; Urbina, Julio A; de Souza, Wanderley

2007-05-15

89

Blockade of tolerance to morphine but not to kappa opioids by a nitric oxide synthase inhibitor.  

PubMed Central

The nitric oxide synthase inhibitor NG-nitro-L-arginine (NO2Arg) blocks morphine tolerance in mice. After implantation of morphine pellets the analgesic response decreases from 100% on the first day to 0% on the third. Coadministration of NO2Arg along with the pellets markedly retards the development of tolerance; 60% of mice are analgesic after 3 days, and 50% of mice are analgesic after 5 days. In a daily injection paradigm the analgesic response to morphine is reduced from 60% to 0% by 5 days. Concomitant administration of morphine along with NO2Arg at doses of 2 mg/kg per day prevents tolerance for 4 weeks. A single NO2Arg dose retards morphine tolerance for several days, and dosing every 4 days is almost as effective as daily NO2Arg. NO2Arg slowly reverses preexisting tolerance over 5 days despite the continued administration of morphine along with NO2Arg. NO2Arg also reduces dependence and reverses previously established dependence. NO2Arg does not prevent tolerance to analgesia mediated by the kappa 1 agonist trans-3,4-dichloro-N-methyl-N-[2-(1-pyrrolindinyl)cyclohexyl]- benzene-acetamide (U50,488H) or the kappa 3 agent naloxone benzoylhydrazone, indicating a selective action of NO in the mechanisms of mu tolerance and dependence. PMID:7685116

Kolesnikov, Y A; Pick, C G; Ciszewska, G; Pasternak, G W

1993-01-01

90

Biological adhesive based on carboxymethyl chitin derivatives and chitin nanofibers.  

PubMed

Novel biological adhesives made from chitin derivatives were prepared and evaluated for their adhesive properties and biocompatibility. Chitin derivatives with acrylic groups, such as 2-hydroxy-3-methacryloyloxypropylated carboxymethyl chitin (HMA-CM-chitin), were synthesized and cured by the addition of an aqueous hydrogen peroxide solution as a radical initiator. The adhesive strength of HMA-CM-chitin increased when it was blended with chitin nanofibers (CNFs) or surface-deacetylated chitin nanofibers (S-DACNFs). HMA-CM-chitin/CNFs or HMA-CM-chitin/S-DACNFs have almost equal adhesive strength compared to that of a commercial cyanoacrylate adhesive. Moreover, quick adhesion and induction of inflammatory cells migration were observed in HMA-CM-chitin/CNF and HMA-CM-chitin/S-DACNF. These findings indicate that the composites prepared in this study are promising materials as new biological adhesives. PMID:25542790

Azuma, Kazuo; Nishihara, Masahiro; Shimizu, Haruki; Itoh, Yoshiki; Takashima, Osamu; Osaki, Tomohiro; Itoh, Norihiko; Imagawa, Tomohiro; Murahata, Yusuke; Tsuka, Takeshi; Izawa, Hironori; Ifuku, Shinsuke; Minami, Saburo; Saimoto, Hiroyuki; Okamoto, Yoshiharu; Morimoto, Minoru

2015-02-01

91

Cell Wall Polysaccharide Synthases Are Located in Detergent-Resistant Membrane Microdomains in Oomycetes ? †  

PubMed Central

The pathways responsible for cell wall polysaccharide biosynthesis are vital in eukaryotic microorganisms. The corresponding synthases are potential targets of inhibitors such as fungicides. Despite their fundamental and economical importance, most polysaccharide synthases are not well characterized, and their molecular mechanisms are poorly understood. With the example of Saprolegnia monoica as a model organism, we show that chitin and (1?3)-?-d-glucan synthases are located in detergent-resistant membrane microdomains (DRMs) in oomycetes, a phylum that comprises some of the most devastating microorganisms in the agriculture and aquaculture industries. Interestingly, no cellulose synthase activity was detected in the DRMs. The purified DRMs exhibited similar biochemical features as lipid rafts from animal, plant, and yeast cells, although they contained some species-specific lipids. This report sheds light on the lipid environment of the (1?3)-?-d-glucan and chitin synthases, as well as on the sterol biosynthetic pathways in oomycetes. The results presented here are consistent with a function of lipid rafts in cell polarization and as platforms for sorting specific sets of proteins targeted to the plasma membrane, such as carbohydrate synthases. The involvement of DRMs in the biosynthesis of major cell wall polysaccharides in eukaryotic microorganisms suggests a function of lipid rafts in hyphal morphogenesis and tip growth. PMID:19201970

Briolay, Anne; Bouzenzana, Jamel; Guichardant, Michel; Deshayes, Christian; Sindt, Nicolas; Bessueille, Laurence; Bulone, Vincent

2009-01-01

92

Heme-Coordinating Inhibitors of Neuronal Nitric Oxide Synthase. Iron-Thioether Coordination is Stabilized by Hydrophobic Contacts Without Increased Inhibitor Potency  

PubMed Central

The heme-thioether ligand interaction often occurs between heme iron and native methionine ligands, but thioether-based heme-coordinating (type II) inhibitors are uncommon due to the difficulty in stabilizing the Fe-S bond. Here, a thioether-based inhibitor (3) of neuronal nitric oxide synthase (nNOS) was designed, and its binding was characterized by spectrophotometry and crystallography. A crystal structure of inhibitor 3 coordinated to heme iron was obtained, representing, to our knowledge, the first crystal structure of a thioether inhibitor complexed to any heme enzyme. A series of related potential inhibitors (4-8) also were evaluated. Compounds 4-8 were all found to be type I (non-heme-coordinating) inhibitors of ferric nNOS, but 4 and 6-8 were found to switch to type II upon heme reduction to the ferrous state, reflecting the higher affinity of thioethers for ferrous heme than for ferric heme. Contrary to what has been widely thought, thioether-heme ligation was found not to increase inhibitor potency, illustrating the intrinsic weakness of the thioether-ferric heme linkage. Subtle changes in the alkyl groups attached to the thioether sulfur caused drastic changes in binding conformation, indicating that hydrophobic contacts play a crucial role in stabilizing the thioether-heme coordination. PMID:20014790

Martell, Jeffrey D.; Li, Huiying; Doukov, Tzanko; Martásek, Pavel; Roman, Linda J.; Soltis, Michael; Poulos, Thomas L.; Silverman, Richard B.

2010-01-01

93

Rationales for cancer chemotherapy with PDMP, a specific inhibitor of glucosylceramide synthase.  

PubMed

A proposed weak point in cancer cells is their need to synthesize novel or rare glucosphingolipids. It is further proposed that cancer patients be treated with a drug that slows the synthesis of glucosylceramide, the precursor of a large family of glucosphingolipids. Experimental data are furnished for chemotherapeutic and biochemical effects of PDMP, an analog of glucosylceramide and its precursor, ceramide. Promising results were obtained in the treatment of mice carrying Ehrlich ascites carcinoma cells and rats carrying C6 glioma cells. PDMP was found to be oxidized by cytochrome P-450, but this process could be blocked in vivo with piperonyl butoxide or cimetidine. A high level of blood glucose was found to elevate the size of rat kidneys and their content of UDP-glucose and its product, glucosylceramide. The excessive growth could be blocked by PDMP, which competes with UDP-glc for binding to glucosylceramide synthase. It is suggested that cancer patients be maintained at a low glucose level in order to slow the synthesis of glucosylceramide by tumor cells. Metabolic changes produced by PDMP in cultured cells, besides a rapid deletion of glucosphingolipids, were accumulation of the precursors (ceramide and sphingosine), loss of protein kinase C, and accumulation of diacylglycerol. It is suggested that many of the cellular changes produced by PDMP, such as loss of cell binding, are owing to existence of glucosylceramide-based "islands" floating in the outer cell surface; the islands may contain growth factor receptors and adhesion factors. An inhibitor that blocks sphingolipid synthesis, such as cycloserine, may prove to be a useful adjuvant for therapy with PDMP. PMID:8086032

Radin, N S

1994-01-01

94

Lipophilic Bisphosphonates as Dual Farnesyl/Geranylgeranyl Diphosphate Synthase Inhibitors: An X-ray and NMR Investigation  

PubMed Central

Considerable effort has focused on the development of selective protein farnesyl transferase (FTase) and protein geranylgeranyl transferase (GGTase) inhibitors as cancer chemotherapeutics. Here, we report a new strategy for anti-cancer therapeutic agents involving inhibition of farnesyl diphosphate synthase (FPPS) and geranylgeranyl diphosphate synthase (GGPPS), the two enzymes upstream of FTase and GGTase, by lipophilic bisphosphonates. Due to dual site targeting and decreased polarity, the compounds have activities far greater than do current bisphosphonate drugs in inhibiting tumor cell growth and invasiveness, both in vitro and in vivo. We explore how these compounds inhibit cell growth, how cell activity can be predicted based on enzyme inhibition data, and, using x-ray diffraction, solid state NMR and isothermal titration calorimetry, we show how these compounds bind to FPPS and/or GGPPS. PMID:19309137

Zhang, Yonghui; Cao, Rong; Yin, Fenglin; Hudock, Michael P.; Guo, Rey-Ting; Krysiak, Kilannin; Mukherjee, Sujoy; Gao, Yi-Gui; Robinson, Howard; Song, Yongcheng; No, Joo Hwan; Bergan, Kyle; Leon, Annette; Cass, Lauren; Goddard, Amanda; Chang, Ting-Kai; Lin, Fu-Yang; Van Beek, Ermond; Papapoulos, Socrates; Wang, Andrew H.-J.; Kubo, Tadahiko; Ochi, Mitsuo; Mukkamala, Dushyant; Oldfield, Eric

2009-01-01

95

Pharmacokinetics and pharmacodynamics of terbogrel, a combined thromboxane A2 receptor and synthase inhibitor, in healthy subjects  

PubMed Central

Aims To characterize the pharmacokinetics of terbogrel, a new combined thromboxane A2 (TxA2) receptor and synthase inhibitor, in healthy human subjects after single or multiple oral administration. Methods Forty-eight healthy male subjects received a single oral dose (10, 25, 50, 100, 150 or 200 mg) of terbogrel or placebo and 32 different subjects received one of the following treatments twice daily for 7 days: 50, 100 or 150 mg terbogrel, placebo, or once-a-day 330 mg acetylsalicylic acid. Results Terbogrel was well tolerated without obvious adverse effects following either a single oral dose or administration over 7 days. Plasma drug concentrations were dose-linear and there was no accumulation over 7 days. There was a dose-dependent blockade of TxA2 receptors and of inhibition of thromboxane synthase activity with values for IC50 of 12 ng ml?1 and 6.7 ng ml?1, respectively. At the highest dose tested (150 mg) there was almost complete inhibition of thomboxane synthase and thromboxane receptor occupancy. Even at trough concentrations, receptor occupancy remained above 80% and thromboxane synthase was still completely inhibited. These two activities were associated with a dose-dependent inhibition of platelet aggregation (>80% at the 150 mg dose of terbogrel) and enhanced prostacyclin production. Conclusions Terbogrel is a potent agent having two distinct, complimentary pharmacodynamic actions, namely inhibition of thromboxane synthase and antagonism of the TxA2 receptor. The antithrombotic effect of terbogrel was dose-dependent and was associated with enhanced prostacyclin production. Terbogrel is an attractive candidate for long-term antithrombotic therapy. PMID:15206991

Guth, Brian D; Narjes, Hans; Schubert, Hans-Dieter; Tanswell, Paul; Riedel, Axel; Nehmiz, Gerhard

2004-01-01

96

BGC 945, a Novel Tumor-Selective Thymidylate Synthase Inhibitor Targeted to A-Folate Receptor-Overexpressing Tumors  

Microsoft Academic Search

BGC 945 is a cyclopenta(g)quinazoline-based, thymidylate synthase inhibitor specifically transported into A-folate recep- tor (A-FR)-overexpressing tumors. Affinity of BGC 945 for the A-FR is 70% of the high-affinity ligand folic acid. In contrast to conventionalantifolates,BGC945haslowaffinityforthewidely expressed reduced-folate carrier (RFC). The Ki for isolated thymidylate synthaseis1.2nmol\\/L andtheIC50forinhibitionof the growth of A-FR-negative mouse L1210 or human A431 cells isf7Mmol\\/L.Incontrast,BGC945ishighlypotentinarangeof A-FR-overexpressing human tumor

David D. Gibbs; Davinder S. Theti; Nadya Wood; Matthew Green; Florence Raynaud; Melanie Valenti; Fraser Mitchell; Vassilios Bavetsias; Elisa Henderson

97

Chitin and chitosan nanofibers: electrospinning of chitin and deacetylation of chitin nanofibers  

Microsoft Academic Search

An electrospinning method was used to fabricate chitin nanofibous matrix for wound dressings. Chitin was depolymerized by gamma irradiation to improve its solubility. The electrospinning of chitin was performed with 1,1,1,3,3,3-hexafluoro-2-propanol (HFIP) as a spinning solvent. Morphology of as-spun and deacetylated chitin (chitosan) nanofibers was investigated by scanning electron microscopy. Although as-spun chitin nanofibers had the broad fiber diameter distribution,

Byung-Moo Min; Sung Won Lee; Jung Nam Lim; Young You; Taek Seung Lee; Pil Hyun Kang; Won Ho Park

2004-01-01

98

Temperature dependent spin crossover in neuronal nitric oxide synthase bound with the heme-coordinating thioether inhibitors  

PubMed Central

A series of L-arginine analogue nitric oxide synthase inhibitors with a thioether tail have been shown to form an Fe-S thioether interaction as evidenced by continuous electron density between the Fe and S atoms. Even so, the Fe-S thioether interaction was found to be far less important for inhibitor binding than the hydrophobic interactions between the alkyl group in the thioether tail and surrounding protein (Martell et al., (2010) J. Am. Chem. Soc. 132, 798). However, among the few thioether inhibitors that showed Fe-S thioether interaction in crystal structures, variations in spin state (high-spin or low-spin) were observed dependent upon the heme iron oxidation state and temperature. Since modern synchrotron X-ray data collection is typically carried out at cryogenic temperatures, we reasoned that some of the discrepancies between cryo-crystal structures and room temperature UV-visible spectroscopy could be the result of temperature dependent spin-state changes. We, therefore, have characterized some of these nNOS-thioether inhibitor complexes in both crystal and solution using EPR and UV-visible absorption spectrometry as a function of temperature and the heme iron redox state. We found that some thioether inhibitors switch from high- to low-spin at lower temperatures similar to the “spin crossover” phenomenon observed in many transition metal complexes. PMID:21534614

Doukov, Tzanko; Li, Huiying; Sharma, Ajay; Martell, Jeffrey D.; Soltis, Michael; Silverman, Richard B.; Poulos, Thomas L.

2011-01-01

99

IF1, a natural inhibitor of mitochondrial ATP synthase, is not essential for the normal growth and breeding of mice  

PubMed Central

IF1 is an endogenous inhibitor protein of mitochondrial ATP synthase. It is evolutionarily conserved throughout all eukaryotes and it has been proposed to play crucial roles in prevention of the wasteful reverse reaction of ATP synthase, in the metabolic shift from oxidative phosphorylation to glycolysis, in the suppression of ROS (reactive oxygen species) generation, in mitochondria morphology and in haem biosynthesis in mitochondria, which leads to anaemia. Here, we report the phenotype of a mouse strain in which IF1 gene was destroyed. Unexpectedly, individuals of this IF1-KO (knockout) mouse strain grew and bred without defect. The general behaviours, blood test results and responses to starvation of the IF1-KO mice were apparently normal. There were no abnormalities in the tissue anatomy or the autophagy. Mitochondria of the IF1-KO mice were normal in morphology, in the content of ATP synthase molecules and in ATP synthesis activity. Thus, IF1 is not an essential protein for mice despite its ubiquitous presence in eukaryotes. PMID:23889209

Nakamura, Junji; Fujikawa, Makoto; Yoshida, Masasuke

2013-01-01

100

The Chitin Connection  

PubMed Central

ABSTRACT Chitin, a polymer of N-acetylglucosamine, is an essential component of the fungal cell wall. Chitosan, a deacetylated form of chitin, is also important in maintaining cell wall integrity and is essential for Cryptococcus neoformans virulence. In their article, Gilbert et al. [N. M. Gilbert, L. G. Baker, C. A. Specht, and J. K. Lodge, mBio 3(1):e00007-12, 2012] demonstrate that the enzyme responsible for chitosan synthesis, chitin deacetylase (CDA), is differentially attached to the cell membrane and wall. Bioactivity is localized to the cell membrane, where it is covalently linked via a glycosylphosphatidylinositol (GPI) anchor. Findings from this study significantly enhance our understanding of cryptococcal cell wall biology. Besides the role of chitin in supporting structural stability, chitin and host enzymes with chitinase activity have an important role in host defense and modifying the inflammatory response. Thus, chitin appears to provide a link between the fungus and host that involves both innate and adaptive immune responses. Recently, there has been increased attention to the role of chitinases in the pathogenesis of allergic inflammation, especially asthma. We review these findings and explore the possible connection between fungal infections, the induction of chitinases, and asthma. PMID:22448043

Goldman, David L.; Vicencio, Alfin G.

2012-01-01

101

Chitin is a necessary component to maintain the barrier function of the peritrophic matrix in the insect midgut.  

PubMed

In most insects, the peritrophic matrix (PM) partitions the midgut into different digestive compartments, and functions as a protective barrier against abrasive particles and microbial infections. In a previous study we demonstrated that certain PM proteins are essential in maintaining the PM's barrier function and establishing a gradient of PM permeability from the anterior to the posterior part of the midgut which facilitates digestion (Agrawal et al., 2014). In this study, we focused on the effects of a reduction in chitin content on PM permeability in larvae of the red flour beetle, Tribolium castaneum. Oral administration of the chitin synthesis inhibitor diflubenzuron (DFB) only partially reduced chitin content of the larval PM even at high concentrations. We observed no nutritional effects, as larval growth was unaffected and neutral lipids were not depleted from the fat body. However, the metamorphic molt was disrupted and the insects died at the pharate pupal stage, presumably due to DFB's effect on cuticle formation. RNAi to knock-down expression of the gene encoding chitin synthase 2 in T. castaneum (TcCHS-2) caused a complete loss of chitin in the PM. Larval growth was significantly reduced, and the fat body was depleted of neutral lipids. In situ PM permeability assays monitoring the distribution of FITC dextrans after DFB exposure or RNAi for TcCHS-2 revealed that PM permeability was increased in both cases. RNAi for TcCHS-2, however, led to a higher permeation of the PM by FITC dextrans than DFB treatment even at high doses. Similar effects were observed when the chitin content was reduced by feeding DFB to adult yellow fever mosquitos, Aedes aegypti. We demonstrate that the presence of chitin is necessary for maintaining the PM's barrier function in insects. It seems that the insecticidal effects of DFB are mediated by the disruption of cuticle synthesis during the metamorphic molt rather than by interfering with larval nutrition. However, as DFB clearly affects PM permeability, it may be suitable to increase the efficiency of pesticides targeting the midgut. PMID:25449129

Kelkenberg, Marco; Odman-Naresh, Jothini; Muthukrishnan, Subbaratnam; Merzendorfer, Hans

2015-01-01

102

Arsenic toxicity induced endothelial dysfunction and dementia: Pharmacological interdiction by histone deacetylase and inducible nitric oxide synthase inhibitors  

SciTech Connect

Arsenic toxicity has been reported to damage all the major organs including the brain and vasculature. Dementia including Alzheimer's disease (AD) and vascular dementia (VaD) are posing greater risk to the world population as it is now increasing at a faster rate. We have investigated the role of sodium butyrate, a selective histone deacetylase (HDAC) inhibitor and aminoguanidine, a selective inducible nitric oxide synthase (iNOS) inhibitor in pharmacological interdiction of arsenic toxicity induced vascular endothelial dysfunction and dementia in rats. Arsenic toxicity was done by administering arsenic drinking water to rats. Morris water-maze (MWM) test was used for assessment of learning and memory. Endothelial function was assessed using student physiograph. Oxidative stress (aortic superoxide anion, serum and brain thiobarbituric acid reactive species, brain glutathione) and nitric oxide levels (serum nitrite/nitrate) were also measured. Arsenic treated rats have shown impairment of endothelial function, learning and memory, reduction in serum nitrite/nitrate and brain GSH levels along with increase in serum and brain TBARS. Sodium butyrate as well as aminoguanidine significantly convalesce arsenic induced impairment of learning, memory, endothelial function, and alterations in various biochemical parameters. It may be concluded that arsenic induces endothelial dysfunction and dementia, whereas, sodium butyrate, a HDAC inhibitor as well as aminoguanidine, a selective iNOS inhibitor may be considered as potential agents for the management of arsenic induced endothelial dysfunction and dementia. - Highlights: • As has induced endothelial dysfunction (Edf) and vascular dementia (VaD). • As has increased oxidative stress, AChE activity and decreased serum NO. • Inhibitors of HDAC and iNOS have attenuated As induced Edf and VaD. • Both the inhibitors have attenuated As induced biochemical changes. • Inhibitor of HDAC and iNOS has shown good potential in As induced VaD.

Sharma, Bhupesh, E-mail: drbhupeshresearch@gmail.com; Sharma, P.M.

2013-11-15

103

L-NAME, a nitric oxide synthase inhibitor, as a potential countermeasure to post-suspension hypotension in rats  

NASA Technical Reports Server (NTRS)

A large number of astronauts returning from spaceflight experience orthostatic hypotension. This hypotension may be due to overproduction of vasodilatory mediators, such as nitric oxide (NO) and prostaglandins. To evaluate the role of the NO synthase inhibitor NG-nitro-L-arginine methyl ester (L-NAME) as a countermeasure against the post-suspension reduction in mean arterial pressure (MAP), we assessed the cardiovascular responses and vascular reactivity to 7-day 30 degrees tail-suspension and a subsequent 6 hr post-suspension period in conscious rats. After a pre-suspension reading, direct MAP and heart rate (HR) were measured daily and every 2 hrs post-suspension. The NO synthase inhibitor L-NAME (20 mg/kg, i.v.), or saline, were administered after the 7th day reading prior to release from suspension and at 2 and 4 hrs post-suspension. At 6 hrs post-suspension, vascular reactivity was assessed. While MAP did not change during the suspension period, it was reduced post-suspension. Heart rate was not significantly altered. L-NAME administration reversed the post-suspension reduction in MAP. In addition, the baroreflex sensitivity for heart rate was modified by L-NAME. Thus, the post-suspension reduction in MAP may be due to overproduction of NO and altered baroreflex activity.

Bayorh, M. A.; Socci, R. R.; Watts, S.; Wang, M.; Eatman, D.; Emmett, N.; Thierry-Palmer, M.

2001-01-01

104

Persistence of effects of nitric oxide synthase inhibitors: comparisons on blood flow and plasma exudation in guinea pig skin.  

PubMed

Plasma protein extravasation has been measured in guinea pig skin using 125I-albumin and blood flow using 133Xenon (133Xe) clearance. The nitric oxide (NO) synthase inhibitors N(G)-nitro-L-arginine methyl ester (L-NAME), N(G)-monomethyl-L-arginine (l-NMMA) and N(G)-nitro-L-arginine (L-NOArg) and the alpha-adrenoceptor agonist, phenylephrine, inhibited bradykinin induced plasma protein extravasation when co-injected with the peptide. The inhibitory effects of L-NAME and L-NOArg lasted for up to 8 and 4 h, respectively, whereas phenylephrine and L-NMMA had no persistent inhibitory effects. When co-injected with 133Xe, L-NAME, L-NMMA, L-NOArg and phenylephrine, but not D-NAME, produced significant reductions in skin blood flow. When injected prior to 133Xe, L-NAME and L-NOArg, but not phenylephrine or L-NMMA, significantly reduced flow. The effect of L-NAME on flow was not significant at 8 h. Thus, although the inhibitory effects of the NO synthase inhibitors on mediator induced plasma protein extravasation show correlations with their effects on blood flow, the persistent effect of L-NAME on exudation appears to extend beyond its effect on flow. PMID:9253959

Perez, A C; Khawaja, A M; Page, C P; Paul, W

1997-07-01

105

Protective effects of a squalene synthase inhibitor, lapaquistat acetate (TAK-475), on statin-induced myotoxicity in guinea pigs.  

PubMed

High-dose statin treatment has been recommended as a primary strategy for aggressive reduction of LDL cholesterol levels and protection against coronary artery disease. The effectiveness of high-dose statins may be limited by their potential for myotoxic side effects. There is currently little known about the molecular mechanisms of statin-induced myotoxicity. Previously we showed that T-91485, an active metabolite of the squalene synthase inhibitor lapaquistat acetate (lapaquistat: a previous name is TAK-475), attenuated statin-induced cytotoxicity in human skeletal muscle cells [Nishimoto, T., Tozawa, R., Amano, Y., Wada, T., Imura, Y., Sugiyama, Y., 2003a. Comparing myotoxic effects of squalene synthase inhibitor, T-91485, and 3-hydroxy-3-methylglutaryl coenzyme A. Biochem. Pharmacol. 66, 2133-2139]. In the current study, we investigated the effects of lapaquistat administration on statin-induced myotoxicity in vivo. Guinea pigs were treated with either high-dose cerivastatin (1 mg/kg) or cerivastatin together with lapaquistat (30 mg/kg) for 14 days. Treatment with cerivastatin alone decreased plasma cholesterol levels by 45% and increased creatine kinase (CK) levels by more than 10-fold (a marker of myotoxicity). The plasma CK levels positively correlated with the severity of skeletal muscle lesions as assessed by histopathology. Co-administration of lapaquistat almost completely prevented the cerivastatin-induced myotoxicity. Administration of mevalonolactone (100 mg/kg b.i.d.) prevented the cerivastatin-induced myotoxicity, confirming that this effect is directly related to HMG-CoA reductase inhibition. These results strongly suggest that cerivastatin-induced myotoxicity is due to depletion of mevalonate derived isoprenoids. In addition, squalene synthase inhibition could potentially be used clinically to prevent statin-induced myopathy. PMID:17599378

Nishimoto, Tomoyuki; Ishikawa, Eiichiro; Anayama, Hisashi; Hamajyo, Hitomi; Nagai, Hirofumi; Hirakata, Masao; Tozawa, Ryuichi

2007-08-15

106

NOpiates: Novel Dual Action Neuronal Nitric Oxide Synthase Inhibitors with ?-Opioid Agonist Activity  

PubMed Central

A novel series of benzimidazole designed multiple ligands (DMLs) with activity at the neuronal nitric oxide synthase (nNOS) enzyme and the ?-opioid receptor was developed. Targeting of the structurally dissimilar heme-containing enzyme and the ?-opioid GPCR was predicated on the modulatory role of nitric oxide on ?-opioid receptor function. Structure–activity relationship studies yielded lead compound 24 with excellent nNOS inhibitory activity (IC50 = 0.44 ?M), selectivity over both endothelial nitric oxide synthase (10-fold) and inducible nitric oxide synthase (125-fold), and potent ?-opioid binding affinity, Ki = 5.4 nM. The functional activity as measured in the cyclic adenosine monosphospate secondary messenger assay resulted in full agonist activity (EC50 = 0.34 ?M). This work represents a novel approach in the development of new analgesics for the treatment of pain. PMID:24900459

2012-01-01

107

NOpiates: Novel Dual Action Neuronal Nitric Oxide Synthase Inhibitors with ?-Opioid Agonist Activity.  

PubMed

A novel series of benzimidazole designed multiple ligands (DMLs) with activity at the neuronal nitric oxide synthase (nNOS) enzyme and the ?-opioid receptor was developed. Targeting of the structurally dissimilar heme-containing enzyme and the ?-opioid GPCR was predicated on the modulatory role of nitric oxide on ?-opioid receptor function. Structure-activity relationship studies yielded lead compound 24 with excellent nNOS inhibitory activity (IC50 = 0.44 ?M), selectivity over both endothelial nitric oxide synthase (10-fold) and inducible nitric oxide synthase (125-fold), and potent ?-opioid binding affinity, K i = 5.4 nM. The functional activity as measured in the cyclic adenosine monosphospate secondary messenger assay resulted in full agonist activity (EC50 = 0.34 ?M). This work represents a novel approach in the development of new analgesics for the treatment of pain. PMID:24900459

Renton, Paul; Green, Brenda; Maddaford, Shawn; Rakhit, Suman; Andrews, John S

2012-03-01

108

Novel 2,4-Disubstituted Pyrimidines as Potent, Selective, and Cell-Permeable Inhibitors of Neuronal Nitric Oxide Synthase  

PubMed Central

Selective inhibition of neuronal nitric oxide synthase (nNOS) is an important therapeutic approach to target neurodegenerative disorders. However, the majority of the nNOS inhibitors developed are arginine mimetics and, therefore, suffer from poor bioavailability. We designed a novel strategy to combine a more pharmacokinetically favorable 2-imidazolylpyrimidine head with promising structural components from previous inhibitors. In conjunction with extensive structure–activity studies, several highly potent and selective inhibitors of nNOS were discovered. X-ray crystallographic analysis reveals that these type II inhibitors utilize the same hydrophobic pocket to gain strong inhibitory potency (13), as well as high isoform selectivity. Interestingly, select compounds from this series (9) showed good permeability and low efflux in a Caco-2 assay, suggesting potential oral bioavailability, and exhibited minimal off-target binding to 50 central nervous system receptors. Furthermore, even with heme-coordinating groups in the molecule, modifying other pharmacophoric fragments minimized undesirable inhibition of cytochrome P450s from human liver microsomes. PMID:25489882

2014-01-01

109

Potent and Selective Double-Headed Thiophene-2-carboximidamide Inhibitors of Neuronal Nitric Oxide Synthase for the Treatment of Melanoma  

PubMed Central

Selective inhibitors of neuronal nitric oxide synthase (nNOS) are regarded as valuable and powerful agents with therapeutic potential for the treatment of chronic neurodegenerative pathologies and human melanoma. Here, we describe a novel hybrid strategy that combines the pharmacokinetically promising thiophene-2-carboximidamide fragment and structural features of our previously reported potent and selective aminopyridine inhibitors. Two inhibitors, 13 and 14, show low nanomolar inhibitory potency (Ki = 5 nM for nNOS) and good isoform selectivities (nNOS over eNOS [440- and 540-fold, respectively] and over iNOS [260- and 340-fold, respectively]). The crystal structures of these nNOS–inhibitor complexes reveal a new hot spot that explains the selectivity of 14 and why converting the secondary to tertiary amine leads to enhanced selectivity. More importantly, these compounds are the first highly potent and selective nNOS inhibitory agents that exhibit excellent in vitro efficacy in melanoma cell lines. PMID:24447275

2015-01-01

110

Novel 2,4-disubstituted pyrimidines as potent, selective, and cell-permeable inhibitors of neuronal nitric oxide synthase.  

PubMed

Selective inhibition of neuronal nitric oxide synthase (nNOS) is an important therapeutic approach to target neurodegenerative disorders. However, the majority of the nNOS inhibitors developed are arginine mimetics and, therefore, suffer from poor bioavailability. We designed a novel strategy to combine a more pharmacokinetically favorable 2-imidazolylpyrimidine head with promising structural components from previous inhibitors. In conjunction with extensive structure-activity studies, several highly potent and selective inhibitors of nNOS were discovered. X-ray crystallographic analysis reveals that these type II inhibitors utilize the same hydrophobic pocket to gain strong inhibitory potency (13), as well as high isoform selectivity. Interestingly, select compounds from this series (9) showed good permeability and low efflux in a Caco-2 assay, suggesting potential oral bioavailability, and exhibited minimal off-target binding to 50 central nervous system receptors. Furthermore, even with heme-coordinating groups in the molecule, modifying other pharmacophoric fragments minimized undesirable inhibition of cytochrome P450s from human liver microsomes. PMID:25489882

Mukherjee, Paramita; Li, Huiying; Sevrioukova, Irina; Chreifi, Georges; Martásek, Pavel; Roman, Linda J; Poulos, Thomas L; Silverman, Richard B

2015-02-12

111

Lipid-lowering effects of TAK-475, a squalene synthase inhibitor, in animal models of familial hypercholesterolemia.  

PubMed

The lipid-lowering effects of 1-[2-[(3R,5S)-1-(3-acetoxy-2,2-dimethylpropyl)-7-chloro-1,2,3,5-tetrahydro-2-oxo-5-(2,3-dimethoxyphenyl)-4,1-benzoxazepine-3-yl] acetyl] piperidin-4-acetic acid (TAK-475), a novel squalene synthase inhibitor, were examined in two models of familial hypercholesterolemia, low-density lipoprotein (LDL) receptor knockout mice and Watanabe heritable hyperlipidemic (WHHL) rabbits. Two weeks of treatment with TAK-475 in a diet admixture (0.02% and 0.07%; approximately 30 and 110 mg/kg/day, respectively) significantly lowered plasma non-high-density lipoprotein (HDL) cholesterol levels by 19% and 41%, respectively, in homozygous LDL receptor knockout mice. The 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitors, simvastatin and atorvastatin (in 0.02% and 0.07% admixtures), also reduced plasma levels of non-HDL cholesterol. In homozygous WHHL rabbits, 4 weeks of treatment with TAK-475 (0.27%; approximately 100 mg/kg/day) lowered plasma total cholesterol, triglyceride and phospholipid levels by 17%, 52% and 26%, respectively. In Triton WR-1339-treated rabbits, TAK-475 inhibited to the same extent the rate of secretion from the liver of the cholesterol, triglyceride and phospholipid components of very-low-density lipoprotein (VLDL). These results suggest that the lipid-lowering effects of TAK-475 in WHHL rabbits are based partially on the inhibition of secretion of VLDL from the liver. TAK-475 had no effect on plasma aspartate aminotransferase and alanine aminotransferase activities. Thus, the squalene synthase inhibitor TAK-475 revealed lipid-lowering effects in both LDL receptor knockout mice and WHHL rabbits. PMID:12679152

Amano, Yuichiro; Nishimoto, Tomoyuki; Tozawa, Ryu ichi; Ishikawa, Eiichiro; Imura, Yoshimi; Sugiyama, Yasuo

2003-04-11

112

Evaluation of substituted triazol-1-yl-pyrimidines as inhibitors of Bacillus anthracis acetohydroxyacid synthase  

Microsoft Academic Search

Acetohydroxyacid synthase (AHAS), a potential target for antimicrobial agents, catalyzes the first common step in the biosynthesis of the branched-chain amino acids. The genes of both catalytic and regulatory subunits of AHAS from Bacillus anthracis (Bantx), a causative agent of anthrax, were cloned, overexpressed in Escherichiacoli, and purified to homogeneity. To develop novel anti-anthracis drugs that inhibit AHAS, a chemical

Vinayakumar Gedi; Kumaresan Jayaraman; Satish Kalme; Hye-Yeon Park; Hae-Chul Park; Im-Joung La; Hoh-Gyu Hahn; Moon-Young Yoon

2010-01-01

113

Combined treatment of ascorbic acid or alpha-tocopherol with dopamine receptor antagonist or nitric oxide synthase inhibitor potentiates cataleptic effect in mice  

Microsoft Academic Search

Rationale  Drugs like haloperidol (Hal) that decrease dopamine (DA) neurotransmission in the striatum induce catalepsy in rodents and\\u000a Parkinson disease-like symptoms in humans. Nitric oxide synthase (NOS) inhibitors interfere with motor activity, disrupting\\u000a rodent exploratory behavior and inducing catalepsy. Catalepsy induced by NOS inhibitors probably involves striatal DA-mediated\\u000a neurotransmission. Antioxidants such as ascorbic acid (vitamin C) and alpha-tocopherol (vitamin E) have

M. Lazzarini; C. Salum; E. A. Del Bel

2005-01-01

114

The NO Donor Sodium Nitroprusside Reverses the Negative Effects on Hepatic Arterial Flow Induced by Endotoxin and the NO Synthase Inhibitor L-NAME  

Microsoft Academic Search

In previous studies we have observed that the nitric oxide synthase inhibitor L-NAME induces a profound deterioration of liver circulation in experimental endotoxemia. Using the same porcine model we now have evaluated the possibility of modulating these effects with the nitric oxide donor sodium nitroprusside. Infusion of endotoxin led to a gradual deterioration of hemodynamic parameters, including liver blood flow.

Y. Gundersen; T. Sætre; T. Scholz; H. Carlsen; H. Kjekshus; O. A. Smiseth; P. Lilleaasen; A. O. Aasen

1996-01-01

115

Deciphering the Genetic Programme Triggering Timely and Spatially-Regulated Chitin Deposition  

PubMed Central

Organ and tissue formation requires a finely tuned temporal and spatial regulation of differentiation programmes. This is necessary to balance sufficient plasticity to undergo morphogenesis with the acquisition of the mature traits needed for physiological activity. Here we addressed this issue by analysing the deposition of the chitinous extracellular matrix of Drosophila, an essential element of the cuticle (skin) and respiratory system (tracheae) in this insect. Chitin deposition requires the activity of the chitin synthase Krotzkopf verkehrt (Kkv). Our data demonstrate that this process equally requires the activity of two other genes, namely expansion (exp) and rebuf (reb). We found that Exp and Reb have interchangeable functions, and in their absence no chitin is produced, in spite of the presence of Kkv. Conversely, when Kkv and Exp/Reb are co-expressed in the ectoderm, they promote chitin deposition, even in tissues normally devoid of this polysaccharide. Therefore, our results indicate that both functions are not only required but also sufficient to trigger chitin accumulation. We show that this mechanism is highly regulated in time and space, ensuring chitin accumulation in the correct tissues and developmental stages. Accordingly, we observed that unregulated chitin deposition disturbs morphogenesis, thus highlighting the need for tight regulation of this process. In summary, here we identify the genetic programme that triggers the timely and spatially regulated deposition of chitin and thus provide new insights into the extracellular matrix maturation required for physiological activity. PMID:25617778

Rotstein, Bárbara; Casali, Andreu; Llimargas, Marta

2015-01-01

116

The crystal structure of spermidine synthase with a multisubstrate adduct inhibitor  

Microsoft Academic Search

Polyamines are essential in all branches of life. Spermidine synthase (putrescine aminopropyltransferase, PAPT) catalyzes the biosynthesis of spermidine, a ubiquitous polyamine. The crystal structure of the PAPT from Thermotoga maritima (TmPAPT) has been solved to 1.5 Angstroms resolution in the presence and absence of AdoDATO (S-adenosyl-1,8-diamino-3-thiooctane), a compound containing both substrate and product moieties. This, the first structure of an

Sergey Korolev; Yoshihiko Ikeguchi; Tatiana Skarina; Steven Beasley; Cheryl Arrowsmith; Alexei Savchenko; Aled Edwards; Andrzej Joachimiak; Anthony E. Pegg

2001-01-01

117

Flavin-Dependent Thymidylate Synthase as a Drug Target for Deadly Microbes: Mutational Study and a Strategy for Inhibitor Design.  

PubMed

The identification of flavin-dependent thymidylate synthase (FDTS) as an essential enzyme and its occurrence in several pathogenic microbes opens opportunities for using FDTS enzyme as an excellent target for new antimicrobial drug discovery. In contrast to the human thymidylate synthase enzyme that utilizes methylene-tetrahydrofolate (CH2H4 folate) for the conversion of dUMP to dTMP, the microbial enzymes utilize an additional non-covalently bound FAD molecule for the hydride transfer from NAD(P)H. The structural and mechanistic differences between the human and microbial enzymes present an attractive opportunity for the design of antimicrobial compounds specific for the pathogens. We have determined the crystal structure of FDTS enzyme in complex with the methyl donor, CH2H4 folate. We describe here the structure of a FDTS mutant and compare it with other FDTS complex structures, including a FDTS-CH2H4 folate complex. We identified a conformational change essential for substrate binding and propose a strategy for the design of FDTS specific inhibitors. PMID:24563811

Mathews, Irimpan I

2013-04-20

118

Effect of nitrovasodilators and inhibitors of nitric oxide synthase on ischaemic and reperfusion function of rat isolated hearts  

PubMed Central

The functional role of the nitric oxide (NO)/guanosine 3?:5?-cyclic monophosphate (cyclic GMP) pathway in experimental myocardial ischaemia and reperfusion was studied in rat isolated hearts.Rat isolated hearts were perfused at constant pressure with Krebs-Henseleit buffer for 25?min (baseline), then made ischaemic by reducing coronary flow to 0.2?ml?min?1 for 25 or 40?min, and reperfused at constant pressure for 25?min. Drugs inhibiting or stimulating the NO/cyclic GMP pathway were infused during the ischaemic phase only. Ischaemic contracture, myocardial cyclic GMP and cyclic AMP levels during ischaemia, and recovery of reperfusion mechanical function were monitored.At baseline, heart rate was 287±12 beats min?1, coronary flow was 12.8±0.6?ml?min?1, left ventricular developed pressure (LVDevP) was 105±4?mmHg and left ventricular end-diastolic pressure 4.6±0.2?mmHg in vehicle-treated hearts (control; n=12). Baseline values were similar in all treatment groups (P>0.05).In normoxic perfused hearts, 1??M NG-nitro-L-arginine (L-NOARG) significantly reduced coronary flow from 13.5±0.2 to 12.1±0.1?ml?min?1 (10%) and LVDevP from 97±1 to 92±1?mmHg (5%; P<0.05, n=5).Ischaemic contracture was 46±2?mmHg, i.e. 44% of LVDevP in control hearts (n=12), unaffected by low concentrations of nitroprusside (1 and 10??M) but reduced to ?30?mmHg (?25%) at higher concentrations (100 or 1000??M; P<0.05 vs control, n=6). Conversely, the NO synthase inhibitor L-NOARG reduced contracture at 1??M to 26±3?mmHg (23%), but increased it to 63±4?mmHg (59%) at 1000??M (n=6). Dobutamine (10??M) exacerbated ischaemic contracture (81±3?mmHg; n=7) and the cyclic GMP analogue Sp-8-(4-p-chlorophenylthio)-3?,5?-monophosphorothioate (Sp-8-pCPT-cGMPS; 10??M) blocked this effect (63±1?mmHg; P<0.05 vs dobutamine alone, n=5).At the end of reperfusion, LVDevP was 58±5?mmHg, i.e. 55% of pre-ischaemic value in control hearts, significantly increased to ?80% by high concentrations of nitroprusside (100 or 1000??M) or L-NOARG at 1??M, while a high concentration of L-NOARG (1000??M) reduced LVDevP to ?35% (P<0.05 vs control; n=6).Ischaemia increased tissue cyclic GMP levels 1.8 fold in control hearts (P<0.05; n=12); nitroprusside at 1??M had no sustained effect, but increased cyclic GMP ?6 fold at 1000??M; L-NOARG (1 or 1000??M) was without effect (n=6). Nitroprusside (1 or 1000??M) marginally increased cyclic AMP levels whereas NO synthase inhibitors had no effect (n=6).In conclusion, the cardioprotective effect of NO donors, but not of low concentrations of NO synthase inhibitors may be due to their ability to elevate cyclic GMP levels. Because myocardial cyclic GMP levels were not affected by low concentrations of NO synthase inhibitors, their beneficial effect on ischaemic and reperfusion function is probably not accompanied by reduced formation of NO and peroxynitrite in this model. PMID:9559900

du Toit, Eugene F; McCarthy, Joy; Miyashiro, Jody; Opie, Lionel H; Brunner, Friedrich

1998-01-01

119

Comparison of nitric oxide synthase inhibitors, phospholipase A2 inhibitor and free radical scavengers as attenuators of opioid withdrawal syndrome.  

PubMed

Chronic morphine-induced withdrawal syndrome after morphine cessation remains a severe obstacle in the clinical treatment of morphine. Previous studies have shown that nitric oxide synthetase (NOS) inhibitors may have therapeutic potential in morphine withdrawal in humans. The mechanisms that underlie expression of morphine-induced withdrawal syndrome are, however, not yet fully understood. Therefore, this study was designed to determine the mechanism of the expression of morphine-induced withdrawal syndrome in mice. Morphine-dependent mice showed marked body weight loss and several withdrawal signs after naloxone challenge. Pretreatment with a NOS inhibitor, such as N-nitro-L-arginine methyl ester (L-NAME) or 7-nitroindazole, but not aminoguanidine, significantly attenuated the expression of morphine-induced withdrawal syndrome. Furthermore, mepacrine (a phospholipase A2 inhibitor) significantly attenuated the morphine-induced withdrawal syndrome in a manner that was different than that with a NOS inhibitor. These results suggest that nNOS and phospholipase A2, which might increase free radicals, play an important role in the expression of morphine-induced withdrawal syndrome. On the contrary, free radical scavengers (including fullerenes, ascorbate-2-phosphate, and DL-alpha-tocopheryl phosphate) attenuated the expression of the morphine-induced withdrawal syndrome. These results indicate that free radicals play an important role in the expression of physical dependence on morphine, and fullerenes could be a potential clinical tool in the relief of morphine withdrawal syndrome. PMID:17989510

Mori, Tomohisa; Ito, Shinobu; Matsubayashi, Kenji; Sawaguchi, Toshiko

2007-12-01

120

Antiproliferative and ultrastructural effects of BPQ-OH, a specific inhibitor of squalene synthase, on Leishmania amazonensis.  

PubMed

Parasites of the Leishmania genus require for the growth and viability the de novo synthesis of specific sterols as such as episterol and 5-dehydroepisterol because cholesterol, which is abundant in their mammalian hosts, does not fulfill the parasite sterol requirements. Squalene synthase catalyzes the first committed step in the sterol biosynthesis and has been studied as a possible target for the treatment of high cholesterol levels in humans. In this work we investigated the antiproliferative and ultrastructural effects induced by 3-(biphenyl-4-yl)-3-hydroxyquinuclidine (BPQ-OH), a specific inhibitor of squalene synthase, on promastigote and amastigote forms of Leishmania amazonensis. BPQ-OH had a potent dose-dependent growth inhibitory effect against promastigotes and amastigotes, with IC(50) values 0.85 and 0.11 microM, respectively. Ultrastructural analysis of the treated parasites revealed several changes in the morphology of promastigote forms. The main ultrastructural change was found in the plasma membrane, which showed signs of disorganization, with the concomitant formation of elaborated structures. We also observed alterations in the mitochondrion-kinetoplast complex such as mitochondrial swelling, rupture of its internal membrane and an abnormal compaction of the kinetoplast. Other alterations included the appearance of multivesicular bodies, myelin-like figures, alterations of the flagellar membrane and presence of parasites with two or more nuclei and kinetoplasts. We conclude that the BPQ-OH was a potent growth inhibitor of L. amazonensis, which led to profound changes of the parasite's ultrastructure and might be a valuable lead compound for the development of novel anti-Leishmania agents. PMID:16198340

Rodrigues, Juliany C F; Urbina, Julio A; de Souza, Wanderley

2005-12-01

121

Genetic rearrangements on the Chlorovirus genome that switch between hyaluronan synthesis and chitin synthesis.  

PubMed

Chlorella viruses or chloroviruses form polysaccharide fibers on the cell wall of host Chlorella cells after infection. Such polysaccharides are either hyaluronan synthesized by virus-encoded hyaluronan synthase (HAS) or chitin synthesized by viral chitin synthase (CHS). Some chloroviruses synthesize both hyaluronan (HA) and chitin simultaneously. To understand the relationship between "HA-synthesizing" and "chitin-synthesizing" viruses, we characterized the CVK2 genomic regions, one flanking chs and the other corresponding to PBCV-1 has and found that on CVK2 DNA, a single ORF (PBCV-1 A330R) was replaced with a 5 kbp region containing chs, ugdh2 (the second gene for UDP-glucose dehydrogenase) and two other ORFs, and that has was replaced with another chs gene. In some chloroviruses, ugdh was lost. These results suggest that chlorovirus types changed from "has viruses" to "chs viruses" or from "chs viruses" to "has viruses" by exchanging the genes. PMID:16112160

Mohammed Ali, Ali Mohammed; Kawasaki, Takeru; Yamada, Takashi

2005-11-10

122

Chitin Biotechnology Applications  

Microsoft Academic Search

This review article describes the current status of the production and consumption of chitin and chitosan, and their current practical applications in biotechnology with some attempted uses. The applications include: 1) cationic agents for polluted waste-water treatment, 2) agricultural materials, 3) food and feed additives, 4) hypocholesterolemic agents, 5) biomedical and pharmaceutical materials, 6) wound-healing materials, 7) blood anticoagulant, antithrombogenic

Shigehiro Hirano

1996-01-01

123

Fatty Acid Synthase Inhibitors Induce Apoptosis in Non-Tumorigenic Melan-A Cells Associated with Inhibition of Mitochondrial Respiration  

PubMed Central

The metabolic enzyme fatty acid synthase (FASN) is responsible for the endogenous synthesis of palmitate, a saturated long-chain fatty acid. In contrast to most normal tissues, a variety of human cancers overexpress FASN. One such cancer is cutaneous melanoma, in which the level of FASN expression is associated with tumor invasion and poor prognosis. We previously reported that two FASN inhibitors, cerulenin and orlistat, induce apoptosis in B16-F10 mouse melanoma cells via the intrinsic apoptosis pathway. Here, we investigated the effects of these inhibitors on non-tumorigenic melan-a cells. Cerulenin and orlistat treatments were found to induce apoptosis and decrease cell proliferation, in addition to inducing the release of mitochondrial cytochrome c and activating caspases-9 and -3. Transfection with FASN siRNA did not result in apoptosis. Mass spectrometry analysis demonstrated that treatment with the FASN inhibitors did not alter either the mitochondrial free fatty acid content or composition. This result suggests that cerulenin- and orlistat-induced apoptosis events are independent of FASN inhibition. Analysis of the energy-linked functions of melan-a mitochondria demonstrated the inhibition of respiration, followed by a significant decrease in mitochondrial membrane potential (??m) and the stimulation of superoxide anion generation. The inhibition of NADH-linked substrate oxidation was approximately 40% and 61% for cerulenin and orlistat treatments, respectively, and the inhibition of succinate oxidation was approximately 46% and 52%, respectively. In contrast, no significant inhibition occurred when respiration was supported by the complex IV substrate N,N,N?,N?-tetramethyl-p-phenylenediamine (TMPD). The protection conferred by the free radical scavenger N-acetyl-cysteine indicates that the FASN inhibitors induced apoptosis through an oxidative stress-associated mechanism. In combination, the present results demonstrate that cerulenin and orlistat induce apoptosis in non-tumorigenic cells via mitochondrial dysfunction, independent of FASN inhibition. PMID:24964211

Rossato, Franco A.; Zecchin, Karina G.; La Guardia, Paolo G.; Ortega, Rose M.; Alberici, Luciane C.; Costa, Rute A. P.; Catharino, Rodrigo R.; Graner, Edgard; Castilho, Roger F.; Vercesi, Aníbal E.

2014-01-01

124

Potentiation of the cytotoxicity of thymidylate synthase (TS) inhibitors by dipyridamole analogues with reduced ?1-acid glycoprotein binding  

PubMed Central

Dipyridamole has been shown to enhance the in vitro activity of antimetabolite anticancer drugs through the inhibition of nucleoside transport. However, the clinical potential of dipyridamole has not been realized because of the avid binding of the drug to the plasma protein ?1-acid glycoprotein (AGP). Dipyridamole analogues that retain potent nucleoside transport inhibitory activity in the presence of AGP are described and their ability to enhance the growth inhibitory and cytotoxic effects of thymidylate synthase (TS) inhibitors has been evaluated. Three dipyridamole analogues (NU3026, NU3059 and NU3060) were shown to enhance the growth inhibitory activity of the TS inhibitor CB3717 and block thymidine rescue in L1210 cells. The extent of potentiation at a fixed analogue concentration (10 ?M) was related to the potency of inhibition of thymidine uptake. A further analogue, NU3076, was identified, which was more potent than dipyridamole with a Ki value for inhibition of thymidine uptake of 0.1 ?M compared to 0.28 ?M for dipyridamole. In marked contrast to dipyridamole, inhibition of thymidine uptake by NU3076 was not significantly affected by the presence of AGP (5 mg ml?1). NU3076 and dipyridamole produced equivalent potentiation of the cytotoxicity of the non-classical antifolate TS inhibitor, nolatrexed, in L1210 cells with both compounds significantly reducing the LC90 by > threefold in the absence of salvageable thymidine. Thymidine rescue of L1210 cells from nolatrexed cytotoxicity was partially blocked by both 1 ?M NU3076 and 1 ?M dipyridamole. NU3076 also caused a significant potentiation of FU cytotoxicity in L1210 cells. These studies demonstrate that nucleoside transport inhibition can be maintained in the absence of AGP binding with the dipyridamole pharmacophore and that such analogues can enhance the cytotoxicity of TS inhibitors. © 1999 Cancer Research Campaign PMID:10468290

Curtin, N J; Bowman, K J; Turner, R N; Huang, B; Loughlin, P J; Calvert, A H; Golding, B T; Griffin, R J; Newell, D R

1999-01-01

125

The design and synthesis of potent and selective inhibitors of Trypanosoma brucei glycogen synthase kinase 3 for the treatment of human african trypanosomiasis.  

PubMed

Glycogen synthase kinase 3 (GSK3) is a genetically validated drug target for human African trypanosomiasis (HAT), also called African sleeping sickness. We report the synthesis and biological evaluation of aminopyrazole derivatives as Trypanosoma brucei GSK3 short inhibitors. Low nanomolar inhibitors, which had high selectivity over the off-target human CDK2 and good selectivity over human GSK3? enzyme, have been prepared. These potent kinase inhibitors demonstrated low micromolar levels of inhibition of the Trypanosoma brucei brucei parasite grown in culture. PMID:25198388

Urich, Robert; Grimaldi, Raffaella; Luksch, Torsten; Frearson, Julie A; Brenk, Ruth; Wyatt, Paul G

2014-09-25

126

The Design and Synthesis of Potent and Selective Inhibitors of Trypanosoma brucei Glycogen Synthase Kinase 3 for the Treatment of Human African Trypanosomiasis  

PubMed Central

Glycogen synthase kinase 3 (GSK3) is a genetically validated drug target for human African trypanosomiasis (HAT), also called African sleeping sickness. We report the synthesis and biological evaluation of aminopyrazole derivatives as Trypanosoma brucei GSK3 short inhibitors. Low nanomolar inhibitors, which had high selectivity over the off-target human CDK2 and good selectivity over human GSK3? enzyme, have been prepared. These potent kinase inhibitors demonstrated low micromolar levels of inhibition of the Trypanosoma brucei brucei parasite grown in culture. PMID:25198388

2014-01-01

127

Marine natural products as inhibitors of cystathionine beta-synthase activity.  

PubMed

A library consisting of characterized marine natural products as well as synthetic derivatives was screened for compounds capable of inhibiting the production of hydrogen sulfide (H2S) by cystathionine beta-synthase (CBS). Eight hits were validated and shown to inhibit CBS activity with IC50 values ranging from 83 to 187?M. The majority of hits came from a series of synthetic polyandrocarpamine derivatives. In addition, a modified fluorogenic probe for H2S detection with improved solubility in aqueous solutions is reported. PMID:25666819

Thorson, Megan K; Van Wagoner, Ryan M; Harper, Mary Kay; Ireland, Chris M; Majtan, Tomas; Kraus, Jan P; Barrios, Amy M

2015-03-01

128

Effects of an endogenous nitric oxide synthase inhibitor on phorbol myristate acetate-induced acute lung injury in rats.  

PubMed

1. In the present study, we determined whether the endogenous nitric oxide (NO) synthase (NOS) inhibitor Nomega-nitro-l-arginine methyl ester (l-NAME) could ameliorate the acute lung injury (ALI) induced by phorbol myristate acetate (PMA) in rat isolated lung. 2. Typical ALI was induced successfully by PMA during 60 min of observation. At 2 micro g/kg, PMA elicited a significant increase in microvascular permeability (measured using the capillary filtration coefficient Kfc), lung weight gain, lung weight/bodyweight ratio, pulmonary arterial pressure (PAP) and protein concentration of bronchoalveolar lavage fluid. 3. Pretreatment with the NOS inhibitor l-NAME (5 mmol/L) significantly attenuated ALI. None of the parameters reflective of lung injury showed significant increase, except for PAP (P < 0.001). The addition of l-arginine (4 mmol/L) blocked the protective effective of l-NAME. Pretreatment with l-arginine exacerbated PMA-induced lung injury. 4. These data suggest that l-NAME significantly ameliorates ALI induced by PMA in rats, indicating that endogenous NO plays a key role in the development of lung oedema in PMA-induced lung injury. PMID:12859432

Lin, Hen I; Chu, Shi Jye; Wang, David; Chen, Hsing I; Hsu, Kang

2003-01-01

129

Secretory leukocyte protease inhibitor suppresses the production of monocyte prostaglandin H synthase-2, prostaglandin E2, and matrix metalloproteinases.  

PubMed Central

Secretory leukocyte protease inhibitor (SLPI) is a serine protease inhibitor found in fluids lining mucosal surfaces. In addition to its primary function as an antiprotease, SLPI may also influence cellular functions associated with enzyme synthesis and retroviral infection. In this study, SLPI was examined for its effect on signaling events involved in the production of matrix metalloproteinases (MMPs) by monocytes. Addition of SLPI before stimulation with concanavalin A or LPS resulted in a significant inhibition of monocyte prostaglandin H synthase-2 (PGHS-2), a pivotal enzyme in the PGE2-cAMP dependent pathway of monocyte MMP synthesis. Suppression of PGHS-2 was detected with 0.1 microg/ml of SLPI with a substantial inhibition at 1 and 10 micro/ml. Attenuation of PGHS-2 by SLPI was accompanied by decreased production of PGE2 resulting in the suppression of interstitial collagenase (MMP-1) and gelatinase B (MMP-9) that was reversed by PGE2 or Bt2cAMP. The inhibitory effect of SLPI was largely independent of its antiprotease activity because SLPI muteins, with significantly lower antiprotease activity, also suppressed the induction of PGHS-2 and MMPs. The inhibitory effects of SLPI did not involve the modulation of monokine production since TNF-alpha and IL-10 were unaffected. These findings demonstrate that SLPI also functions as a potent antiinflammatory agent by interfering with the signal transduction pathway leading to monocyte MMP production. PMID:9062347

Zhang, Y; DeWitt, D L; McNeely, T B; Wahl, S M; Wahl, L M

1997-01-01

130

Catechol-based substrates of chalcone synthase as a scaffold for novel inhibitors of PqsD.  

PubMed

A new strategy for treating Pseudomonas aeruginosa infections could be disrupting the Pseudomonas Quinolone Signal (PQS) quorum sensing (QS) system. The goal is to impair communication among the cells and, hence, reduce the expression of virulence factors and the formation of biofilms. PqsD is an essential enzyme for the synthesis of PQS and shares some features with chalcone synthase (CHS2), an enzyme expressed in Medicago sativa. Both proteins are quite similar concerning the size of the active site, the catalytic residues and the electrostatic surface potential at the entrance of the substrate tunnel. Hence, we evaluated selected substrates of the vegetable enzyme as potential inhibitors of the bacterial protein. This similarity-guided approach led to the identification of a new class of PqsD inhibitors having a catechol structure as an essential feature for activity, a saturated linker with two or more carbons and an ester moiety bearing bulky substituents. The developed compounds showed PqsD inhibition with IC50 values in the single-digit micromolar range. The binding mode of these compounds was investigated by Surface Plasmon Resonance (SPR) experiments revealing that their interaction with the protein is not influenced by the presence of the anthranilic acid bound to active site cysteine. Importantly, some compounds reduced the signal molecule production in cellulo. PMID:25437621

Allegretta, Giuseppe; Weidel, Elisabeth; Empting, Martin; Hartmann, Rolf W

2015-01-27

131

Chitin synthesis in chlorovirus CVK2-infected chlorella cells.  

PubMed

Hyaluronan synthesis in chlorovirus PBCV-1-infected Chlorella cells was previously reported (DeAngelis et al., 1997). In contrast, we report here on the detection, characterization, and expression of a gene for chitin synthase (chs) encoded by chlorovirus CVK2 isolated in Kyoto, Japan. The CVK2 chs gene encoding an open reading frame of 516 aa was expressed as early as 10 min postinfection (p.i.), peaked at 20-40 min p.i., and disappeared at 120-180 min p.i. The chitin polysaccharide began to accumulate as chitinase-sensitive, hair-like fibers on the outside of the virus-infected Chlorella cell wall by 30 min p.i. All chloroviruses without the gene for hyaluronan synthase (has) alternatively contained the chs gene, suggesting the importance of polysaccharide production in the course of virus infection. A few chloroviruses possessed both the chs and has genes and produced chitin and hyaluronan simultaneously. Polysaccharide accumulation on the algal surface may protect virus-infected algae from uptake by other organisms, such as protozoa. Since CVK2 was reported to encode two chitinases and one chitosanase, CVK2 is a very peculiar virus that encodes enzymes required for both the synthesis and the degradation of chitin materials. PMID:12429521

Kawasaki, Takeru; Tanaka, Masahiro; Fujie, Makoto; Usami, Shoji; Sakai, Kazuo; Yamada, Takashi

2002-10-10

132

Inducible Nitric Oxide Synthase Inhibitor SD-3651 Reduces Proteinuria in MRL/lpr Mice Deficient in the NOS2 Gene  

PubMed Central

Several studies have demonstrated the effectiveness of arginine analog nitric oxide synthase (NOS) inhibitor therapy in preventing and treating murine lupus nephritis. However, MRL/MpJ-FASlpr (MRL/lpr) mice lacking a functional NOS2 (inducible NOS [iNOS]) gene (NOS2?/?) develop proliferative glomerulonephritis in a fashion similar to their wild-type (wt) littermates. This finding suggests that the effect of arginine analog NOS inhibitors is through a non-iNOS–mediated mechanism. This study was designed to address this hypothesis. NOS2?/? mice were given either vehicle or a NOS inhibitor (SD-3651) to determine if pharmacological NOS inhibition prevented glomerulonephritis, using wt mice as positive controls. Urine was collected fortnightly to measure albumin. At the time of full disease expression in wt mice, all mice were killed, and renal tissue was examined for light, immunofluorescence, and electron microscopic evidence of disease. Serum was analyzed for anti–double-stranded DNA antibody production. NOS2?/? mice had higher serum anti–double-stranded DNA antibody antibody levels than those of wt mice. SD-3651 therapy reduced proteinuria, glomerular immunoglobulin G deposition, and electron microscopic evidence of podocytopathy and endothelial cell swelling without affecting proliferative lesions by light microscopy. These studies confirm that genetic iNOS deficiency alone is insufficient to prevent proliferative glomerulonephritis and suggest that iNOS activity may inhibit autoantibody production. These results also suggest that SD-3651 therapy acts via a non–iNOS-mediated mechanism to prevent endothelial cell and podocyte pathology. Studies that elucidate this mechanism could provide a useful drug target for the treatment of nephritis. PMID:18797415

Njoku, Chinedu; Self, Sally E.; Ruiz, Philip; Hofbauer, Ann F.; Gilkeson, Gary S.; Oates, Jim C.

2009-01-01

133

Glycogen synthase kinase 3 inhibitors in the next horizon for Alzheimer's disease treatment.  

PubMed

Glycogen synthase kinase 3 (GSK-3), a proline/serine protein kinase ubiquitously expressed and involved in many cellular signaling pathways, plays a key role in the pathogenesis of Alzheimer's disease (AD) being probably the link between ?-amyloid and tau pathology. A great effort has recently been done in the discovery and development of different new molecules, of synthetic and natural origin, able to inhibit this enzyme, and several kinetics mechanisms of binding have been described. The small molecule called tideglusib belonging to the thiadiazolidindione family is currently on phase IIb clinical trials for AD. The potential risks and benefits of this new kind of disease modifying drugs for the future therapy of AD are discussed in this paper. PMID:21760986

Martinez, Ana; Gil, Carmen; Perez, Daniel I

2011-01-01

134

Glycogen Synthase Kinase 3 Inhibitors in the Next Horizon for Alzheimer's Disease Treatment  

PubMed Central

Glycogen synthase kinase 3 (GSK-3), a proline/serine protein kinase ubiquitously expressed and involved in many cellular signaling pathways, plays a key role in the pathogenesis of Alzheimer's disease (AD) being probably the link between ?-amyloid and tau pathology. A great effort has recently been done in the discovery and development of different new molecules, of synthetic and natural origin, able to inhibit this enzyme, and several kinetics mechanisms of binding have been described. The small molecule called tideglusib belonging to the thiadiazolidindione family is currently on phase IIb clinical trials for AD. The potential risks and benefits of this new kind of disease modifying drugs for the future therapy of AD are discussed in this paper. PMID:21760986

Martinez, Ana; Gil, Carmen; Perez, Daniel I.

2011-01-01

135

In silico deconstruction of ATP-competitive inhibitors of glycogen synthase kinase-3?.  

PubMed

Fragment-based methods have emerged in the last two decades as alternatives to traditional high throughput screenings for the identification of chemical starting points in drug discovery. One arguable yet popular assumption about fragment-based design is that the fragment binding mode remains conserved upon chemical expansion. For instance, the question of the binding conservation upon fragmentation of a molecule is still unclear. A number of papers have challenged this hypothesis by means of experimental techniques, with controversial results, "underlining" the idea that a simple generalization, maybe, is not possible. From a computational standpoint, the issue has been rarely addressed and mostly to test novel protocols on limited data sets. To fill this gap, we here report on a computational retrospective study concerned with the in silico deconstruction of leadlike compounds, active on the pharmaceutically relevant enzyme glycogen synthase kinase-3?. PMID:23198830

Bisignano, Paola; Lambruschini, Chiara; Bicego, Manuele; Murino, Vittorio; Favia, Angelo D; Cavalli, Andrea

2012-12-21

136

Chitin Synthesis in Chlorovirus CVK2Infected Chlorella Cells  

Microsoft Academic Search

Hyaluronan synthesis in chlorovirus PBCV-1-infected Chlorella cells was previously reported (DeAngelis et al., 1997). In contrast, we report here on the detection, characterization, and expression of a gene for chitin synthase (chs) encoded by chlorovirus CVK2 isolated in Kyoto, Japan. The CVK2 chs gene encoding an open reading frame of 516 aa was expressed as early as 10 min postinfection

Takeru Kawasaki; Masahiro Tanaka; Makoto Fujie; Shoji Usami; Kazuo Sakai; Takashi Yamada

2002-01-01

137

Inhibition of stimulated ascorbic acid and luteinizing hormone-releasing hormone release by nitric oxide synthase or guanyl cyclase inhibitors.  

PubMed

Ascorbic acid (AA), an antioxidant, is present in high concentrations in the hypothalamus. Previously, we have shown that AA inhibited stimulated release of luteinizing hormone-releasing hormone (LHRH) from medial basal hypothalami in vitro. We have also demonstrated that cell membrane depolarization by high [K(+)] media-induced AA release that is blocked by N(G)-mono-methyl-L-arginine, a competitive inhibitor of nitric oxide synthase (NOS), indicating that the release process is mediated by NO. The release of LHRH is also mediated by NO. We hypothesized that AA is a co-transmitter released with classical transmitters from synaptic vesicles that acts to reduce chemically the NO formed, thereby providing feed-forward inhibitory control over LHRH release. Because NO acts by activating guanylyl cyclase (GC) resulting in production of cGMP, in the present investigation we studied the effects of an NOS inhibitor LY 83583 and GC inhibitor, O.D.Q. to further characterize the role of NO in high [K(+)]-induced AA and LHRH release. Medial basal hypothalami were incubated in 0.5 ml of Krebs-Ringer Bicarbonate buffer or medium containing increased potassium [K(+) = 56 mM] for 1 hr or combinations of high [K(+)] + LY 83583 or O.D.Q. for 1 hr. AA and LHRH released into the incubation medium were measured by high-pressure liquid chromatography and radioimmunoassay, respectively. Cell membrane depolarization with high [K(+)] produced a significant increase in both AA and LHRH release. A combination of high [K(+)] + LY 83583 or high [K(+)] + O.D.Q. decreased basal AA and completely blocked high [K(+)]-induced AA and LHRH release. As in the case of high [K(+)], LHRH release induced by the excitatory amino acid N-methyl-D-aspartic acid (NMDA) was blocked by both the inhibitors. NMDA alone failed to alter AA release, but the combined presence of NMDA and the inhibitors totally blocked AA release. Because LY 83583 and O.D.Q. were shown to inhibit NOS and soluble GC, respectively, the data demonstrate that basal and high [K(+)]-induced AA and high [K(+)] and NMDA-stimulated LHRH release were mediated by NO by its activation of GC and consequent generation of cGMP. PMID:14709779

Karanth, Sharada; Yu, Wen H; Mastronardi, Claudio A; McCann, Samuel M

2004-01-01

138

Ozagrel hydrochloride, a selective thromboxane A2 synthase inhibitor, alleviates liver injury induced by acetaminophen overdose in mice  

PubMed Central

Background Overdosed acetaminophen (paracetamol, N-acetyl-p-aminophenol; APAP) causes severe liver injury. We examined the effects of ozagrel, a selective thromboxane A2 (TXA2) synthase inhibitor, on liver injury induced by APAP overdose in mice. Methods Hepatotoxicity was induced to ICR male mice by an intraperitoneal injection with APAP (330 mg/kg). The effects of ozagrel (200 mg/kg) treatment 30 min after the APAP injection were evaluated with mortality, serum alanine aminotransferase (ALT) levels and hepatic changes, including histopathology, DNA fragmentation, mRNA expression and total glutathione contents. The impact of ozagrel (0.001-1 mg/mL) on cytochrome P450 2E1 (CYP2E1) activity in mouse hepatic microsome was examined. RLC-16 cells, a rat hepatocytes cell line, were exposed to 0.25 mM N-acetyl-p-benzoquinone imine (NAPQI), a hepatotoxic metabolite of APAP. In this model, the cytoprotective effects of ozagrel (1–100 muM) were evaluated by the WST-1 cell viability assay. Results Ozagel treatment significantly attenuated higher mortality, elevated serum alanine aminotransferase levels, excessive hepatic centrilobular necrosis, hemorrhaging and DNA fragmentation, as well as increase in plasma 2,3-dinor thromboxane B2 levels induced by APAP injection. Ozagrel also inhibited the hepatic expression of cell death-related mRNAs induced by APAP, such as jun oncogene, FBJ osteosarcoma oncogene (fos) and C/EBP homologous protein (chop), but did not suppress B-cell lymphoma 2-like protein11 (bim) expression and hepatic total glutathione depletion. These results show ozagrel can inhibit not all hepatic changes but can reduce the hepatic necrosis. Ozagrel had little impact on CYP2E1 activity involving the NAPQI production. In addition, ozagrel significantly attenuated cell injury induced by NAPQI in RLC-16. Conclusions We demonstrate that the TXA2 synthase inhibitor, ozagrel, dramatically alleviates liver injury induced by APAP in mice, and suggest that it is a promising therapeutic candidate for the treatment of APAP-induced liver injury. PMID:23363429

2013-01-01

139

Effect of simvastatin on endothelium-dependent vaso- relaxation and endogenous nitric oxide synthase inhibitor  

Microsoft Academic Search

AIM: To investigate the effect of simvastatin on endothelium-dependent vasorelaxation and endogenous nitric oxide synthesis inhibitor asymmetric dimethylarginine (ADMA) in rats and cultured ECV304 cells. METHODS: Endothe- lial injury was induced by a single injection of low density lipoprotein (LDL) (4 mg\\/kg, 48 h) in rats or incubation with LDL (300 mg\\/L) or oxidative-modified LDL (100 mg\\/L) in cultured ECV304

Jun-lin JIANG; De-jian JIANG; Yu-hai TANG; Nian-sheng LI; Han-wu DENG; Yuan-jian LI

140

Effect of nitric oxide synthase inhibitor and NMDA receptor antagonist on the development of nicotine sensitization of nucleus accumbens dopamine release: An in vivo microdialysis study  

Microsoft Academic Search

We have previously found that the neuronal nitric oxide synthase inhibitor N-nitro-l-arginine (l-NNA) and the noncompetitive N-methyl-d-aspartate (NMDA) receptor antagonist MK-801 prevent behavioral sensitization to nicotine. This study aimed to investigate the effect of l-NNA and MK-801 on a neurochemical component of nicotine sensitization by evaluating the effect of the drugs on nicotine sensitization of nucleus accumbens dopamine (DA) release.

Soo Kyung Hong; In Soon Jung; Seong Ae Bang; Sang Eun Kim

2006-01-01

141

The crystal structure of spermidine synthase with a multisubstrate adduct inhibitor.  

SciTech Connect

Polyamines are essential in all branches of life. Spermidine synthase (putrescine aminopropyltransferase, PAPT) catalyzes the biosynthesis of spermidine, a ubiquitous polyamine. The crystal structure of the PAPT from Thermotoga maritima (TmPAPT) has been solved to 1.5 Angstroms resolution in the presence and absence of AdoDATO (S-adenosyl-1,8-diamino-3-thiooctane), a compound containing both substrate and product moieties. This, the first structure of an aminopropyltransferase, reveals deep cavities for binding substrate and cofactor, and a loop that envelops the active site. The AdoDATO binding site is lined with residues conserved in PAPT enzymes from bacteria to humans, suggesting a universal catalytic mechanism. Other conserved residues act sterically to provide a structural basis for polyamine specificity. The enzyme is tetrameric; each monomer consists of a C-terminal domain with a Rossmann-like fold and an N-terminal {beta}-stranded domain. The tetramer is assembled using a novel barrel-type oligomerization motif.

Korolev, S.; Ikeguchi, Y.; Skarina, T.; Beasley, S.; Arrowsmith, C.; Edwards, A.; Joachimiak, A.; Pegg, A. E.; Savchenko, A.; Pennsylvania State Univ. Coll. of Medicine; Milton S. Hershey Medical Center; Banting and Best Department of Medical Research; Univ. of Health Network

2002-01-01

142

Morlin, an inhibitor of cortical microtubule dynamics and cellulose synthase movement  

PubMed Central

Morlin (7-ethoxy-4-methyl chromen-2-one) was discovered in a screen of 20,000 compounds for small molecules that cause altered cell morphology resulting in swollen root phenotype in Arabidopsis. Live-cell imaging of fluorescently labeled cellulose synthase (CESA) and microtubules showed that morlin acts on the cortical microtubules and alters the movement of CESA. Morlin caused a novel syndrome of cytoskeletal defects, characterized by cortical array reorientation and compromised rates of both microtubule elongation and shrinking. Formation of shorter and more bundled microtubules and detachment from the cell membrane resulted when GFP::MAP4-MBP was used to visualize microtubules during morlin treatment. Cytoskeletal effects were accompanied by a reduction in the velocity and redistribution of CESA complexes labeled with YFP::CESA6 at the cell cortex. Morlin caused no inhibition of mouse myoblast, bacterial or fungal cell proliferation at concentrations that inhibit plant cell growth. By contrast, morlin stimulated microtubule disassembly in cultured hippocampal neurons but had no significant effect on cell viability. Thus, morlin appears to be a useful new probe of the mechanisms that regulate microtubule cortical array organization and its functional interaction with CESA. PMID:17389408

DeBolt, Seth; Gutierrez, Ryan; Ehrhardt, David W.; Melo, Carlos V.; Ross, Loretta; Cutler, Sean R.; Somerville, Christopher; Bonetta, Dario

2007-01-01

143

A Fatal Combination: A Thymidylate Synthase Inhibitor with DNA Damaging Activity  

PubMed Central

2?-deoxy-5-ethynyluridine (EdU) has been previously shown to be a cell poison whose toxicity depends on the particular cell line. The reason is not known. Our data indicates that different efficiency of EdU incorporation plays an important role. The EdU-mediated toxicity was elevated by the inhibition of 2?-deoxythymidine 5?-monophosphate synthesis. EdU incorporation resulted in abnormalities of the cell cycle including the slowdown of the S phase and a decrease in DNA synthesis. The slowdown but not the cessation of the first cell division after EdU administration was observed in all of the tested cell lines. In HeLa cells, a 10 ?M EdU concentration led to the cell death in the 100% of cells probably due to the activation of an intra S phase checkpoint in the subsequent S phase. Our data also indicates that this EdU concentration induces interstrand DNA crosslinks in HeLa cells. We suppose that these crosslinks are the primary DNA damage resulting in cell death. According to our results, the EdU-mediated toxicity is further increased by the inhibition of thymidylate synthase by EdU itself at its higher concentrations. PMID:25671308

Ligasová, Anna; Strunin, Dmytro; Friedecký, David; Adam, Tomáš; Koberna, Karel

2015-01-01

144

Allosteric Inhibitors at the Heterodimer Interface of Imidazole Glycerol Phosphate Synthase  

NASA Astrophysics Data System (ADS)

Imidazole glycerol phosphate synthase (IGPS) from Thermotoga maritima is a heterodimeric enzyme composed of the HisH and HisF proteins. It is attractive as a pathological target since it is absent in mammals but found in plant and opportunistic human pathogens. IGPS was experimentally determined to be a V-type allosteric enzyme that is involved in an essential biosynthetic pathway of microorganisms. The enzyme catalyzes the hydrolysis of glutamine to form NH3 in the HisH protein, followed by cyclization of NH3 with N'-[(5'-phosphoribulosyl)imino]-5-aminoimidazole-4-carboxamide-ribonucleotide (PRFAR) in the HisF subunit, forming imidazole glycerol phosphate (IGP) and 5-aminoimidazole-4-carboxamide ribotide (AICAR) that enter the histidine and purine biosynthetic pathways. Allosteric motions induced upon the binding of the effector PRFAR to HisF propagate through the non-covalent HisH/HisF interface and synchronize catalytic activity at the two distant active sites. However, the nature of the allosteric pathway and the feasibility of manipulating signal transduction by using allosteric drug-like molecules remain to be established. Molecular docking studies of commercial drugs at the HisH/HisF interface were used to identify stable candidates with a potential allosteric effect on the reaction mechanism. Molecular dynamic simulations and calculations of NMR chemical shifts were combined to elucidate the allosteric pathway of IGPS.

Snoeberger, Ning-Shiuan Nicole

145

Internalization and stability of a thymidylate synthase Peptide inhibitor in ovarian cancer cells.  

PubMed

Information on the cellular internalization and stability of the ovarian cancer cell growth inhibitor peptide, LSCQLYQR (LR), is vital for lead optimization. Ad-hoc-synthesized LR/fluorescent-probe conjugates were used to monitor the internalization of the peptide. Mass spectrometry was used to identify adducts resulting from the thiol reactivity of the cysteine residue in LR. A mechanistic model is proposed to explain the observed change in intracellular peptide amount over time. Structural modifications can be foreseen to improve the peptide stability. PMID:25353379

Cannazza, Giuseppe; Cazzato, Addolorata Stefania; Marraccini, Chiara; Pavesi, Giorgia; Pirondi, Silvia; Guerrini, Remo; Pelà, Michela; Frassineti, Chiara; Ferrari, Stefania; Marverti, Gaetano; Ponterini, Glauco; Costi, Maria Paola

2014-12-26

146

Low protein diet exacerbates NO synthase inhibitor-induced hypertension in rats  

Microsoft Academic Search

Background. A low-protein diet (LPD) may suppress nitric oxide (NO) production through the reduction of L-arginine intake, and possibly\\u000a exacerbates hypertension under certain conditions. We examined the effect of LPD on blood pressure in rats that received a\\u000a low dose of an inhibitor of nitric oxide synthesis.\\u000a \\u000a \\u000a Methods. Were compared tail-cuff pressure (TCP), urinary nitrite and nitrate (NO2\\/NO3) excretion, and

Hiromichi Kumagai; Yoshiko Kikuchi; Noriko Kumeta; Masato Kimura; Toshio Sakai

1999-01-01

147

Design and syntheses of novel phthalazin-1(2H)-one derivatives as acetohydroxyacid synthase inhibitors.  

PubMed

A series of 2-substituted-8-(4,6-dimethoxypyrimidin-2-yloxy)-4-methylphthalazin-1-one derivatives, 7a-7w, were designed via an ortho-substituent cyclization strategy to discover a new herbicidal lead structure. These compounds were synthesized by a seven-step route using 3-hydroxy-acetophenone as a starting material. Determination of the Ki values against wild-type A. thaliana acetohydroxyacid synthase (AHAS) (EC 4.1.3.18) indicated that some of the compounds displayed good enzyme inhibition activity comparable to that of KIH-6127. The further preliminary bioassay data on weeds showed that the synthesized compounds exhibited typical injury symptoms of AHAS-inhibiting herbicides, and some of them showed broad-spectrum and high herbicidal activities in postemergence treatments against Echinochloa crusgalli, Digitaria sanguinalis, Setaria viridis, Brassica juncea, Amaranthus retroflexus, and Chenopodium album at an application rate of 150 g ai/ha. To our knowledge, this is the first report of methylphthalazin-1-one derivatives as AHAS inhibitors. PMID:17117801

Li, Yuan-Xiang; Luo, Yan-Ping; Xi, Zhen; Niu, Congwei; He, Yan-Zhen; Yang, Guang-Fu

2006-11-29

148

Effect of nitric oxide synthase inhibitor L-NAME on fear extinction in rats: a task-dependent effect.  

PubMed

There is increasing evidence that nitric oxide may be involved in learning and memory. However, there remain comparatively few studies that have explored the relationship between nitric oxide signaling and fear extinction, an inhibitory learning model. In the present study, we tested the effects of nitric oxide synthase inhibitor l-NAME on three tone fear extinction tasks in rats. In task 1, rats received fear conditioning, extinction training and extinction test in the same context (AAA design). In task 2, rats received fear conditioning in context A, extinction training in context B and extinction test in context A (ABA design). In task 3, rats received fear conditioning in context A, extinction training and extinction test in context B (ABB design). l-NAME (10, 20 and 40 mg/kg) was injected intraperitoneally 30 min prior to extinction training in each task. Percent of time spent freezing was used to measure conditioned fear response. We found that l-NAME administrations had no effect on freezing in task 1 and 2 but produced a dose-dependent increase in task 3. Further results indicated that the increased freezing in task 3 was not attributed to state-dependency effects or nonspecific changes of locomotor activity that followed l-NAME injection. These results showed that l-NAME produced a task-dependent impairment of fear extinction, and implied that nitric oxide signaling was involved in memory process of certain extinction tasks. PMID:24792396

Luo, Huaiqing; Han, Li; Tian, Shaowen

2014-06-20

149

Discovery of a Novel Class of Orally Active Antifungal ?-1,3-d-Glucan Synthase Inhibitors?  

PubMed Central

The echinocandins are a class of semisynthetic natural products that target ?-1,3-glucan synthase (GS). Their proven clinical efficacy combined with minimal safety issues has made the echinocandins an important asset in the management of fungal infection in a variety of patient populations. However, the echinocandins are delivered only parenterally. A screen for antifungal bioactivities combined with mechanism-of-action studies identified a class of piperazinyl-pyridazinones that target GS. The compounds exhibited in vitro activity comparable, and in some cases superior, to that of the echinocandins. The compounds inhibit GS in vitro, and there was a strong correlation between enzyme inhibition and in vitro antifungal activity. In addition, like the echinocandins, the compounds caused a leakage of cytoplasmic contents from yeast and produced a morphological response in molds characteristic of GS inhibitors. Spontaneous mutants of Saccharomyces cerevisiae with reduced susceptibility to the piperazinyl-pyridazinones had substitutions in FKS1. The sites of these substitutions were distinct from those conferring resistance to echinocandins; likewise, echinocandin-resistant isolates remained susceptible to the test compounds. Finally, we present efficacy and pharmacokinetic data on an example of the piperazinyl-pyridazinone compounds that demonstrated efficacy in a murine model of Candida glabrata infection. PMID:21844320

Walker, Scott S.; Xu, Yiming; Triantafyllou, Ilias; Waldman, Michelle F.; Mendrick, Cara; Brown, Nathaniel; Mann, Paul; Chau, Andrew; Patel, Reena; Bauman, Nicholas; Norris, Christine; Antonacci, Barry; Gurnani, Maya; Cacciapuoti, Anthony; McNicholas, Paul M.; Wainhaus, Samuel; Herr, R. Jason; Kuang, Rongze; Aslanian, Robert G.; Ting, Pauline C.; Black, Todd A.

2011-01-01

150

Spontaneous rearrangement of aminoalkylisothioureas into mercaptoalkylguanidines, a novel class of nitric oxide synthase inhibitors with selectivity towards the inducible isoform.  

PubMed Central

1. The generation of nitric oxide (NO) from L-arginine by NO synthases (NOS) can be inhibited by guanidines, amidines and S-alkylisothioureas. Unlike most L-arginine based inhibitors, however, some guanidines and S-alkylisothioureas, in particular aminoethylisothiourea (AETU), show selectivity towards the inducible isoform (iNOS) over the constitutive isoforms (endothelial, ecNOS and brain isoform, bNOS) and so may be of therapeutic benefit. In the present study we have investigated the effects of AETU and other aminoalkylisothioureas on the activities of iNOS, ecNOS and bNOS. 2. AETU, aminopropylisothiourea (APTU) and their derivatives containing alkyl substituents on one of the amidino nitrogens, potently inhibit nitrite formation by immunostimulated J774 macrophages (a model of iNOS activity) with EC50 values ranging from 6-30 microM (EC50 values for NG-methyl-L-arginine (L-NMA) and NG-nitro-L-arginine were 159 and > 1000 microM, respectively). The inhibitory effects of these aminoalkylisothioureas (AATUs) were attentuated by L-arginine in the incubation medium, indicating that these agents may complete with L-arginine for its binding site on NOS. 3. The above AATUs undergo chemical conversion in neutral or basic solution (pH 7 or above) as indicated by (1) the disappearance of AATUs from solution as measured by h.p.l.c., (2) the generation of free thiols not previously present and (3) the isolation of species (as picrate and flavianate salts) from neutral or basic solutions of AATUs that are different from those obtained from acid solutions. 4. Mercaptoalkylguanidines (MAGs) were prepared and shown to be potent inhibitors of iNOS activity with EC50s comparable to those of their isomeric AATUs. 5. These findings suggest that certain AATUs exert their potent inhibitory effects through intramolecular rearrangement to mercaptoalkylguanidines (MAGs) at physiological pH. Those AATUs not capable of such rearrangement do not exhibit the same degree of inhibition of iNOS. 6. In contrast to their potent effects on iNOS, some AATUs and MAGs were 20-100 times weaker than NG-methyl-L-arginine and NG-nitro-L-arginine as inhibitors of ecNOS as assessed by their effects on the conversion of L-arginine to L-citrulline in homogenates of bovine endothelial cells and by their pressor effects in anaesthetized rats. Thus mercaptoalkylguanidines represent a new class of NOS inhibitors with preference towards iNOS. 7. AETU and mercaptoethylguanidine (MEG), when given as infusions, gave slight decreases in MAP in control rats. However, infusions of AETU or MEG to endotoxin-treated rats caused an increase in MAP and restored 80% of the endotoxin-induced fall in MAP. 8. High doses of MEG (30-60 mg kg-1) caused a decrease in MAP of normal rats. This depressor effect may be a consequence of the in vivo oxidation of MEG to the disulphide, guanidinoethyldisulphide (GED), which caused pronounced, transient hypotensive responses in anaesthetized rats and caused endothelium-independent vasodilator responses in precontracted rat aortic rings in vitro. 9. In some cases, slight differences were observed in the activities of AATUs and the corresponding MAGs. These may be explained by the formation of other species from AATUs in physiological media. For example, AETU can give rise to small amounts of the potent ecNOS inhibitor, 2-aminothiazoline, in addition to MEG. This may account for the differences in the in vitro and in vivo effects of AETU and MEG. 10. In conclusion, the in vitro and in vivo effects of AETU and related aminoalkylisothioureas can be explained in terms of their intramolecular rearrangement to generate mercaptoalkylguanidines, a novel class of selective inhibitors of iNOS. PMID:8646406

Southan, G. J.; Zingarelli, B.; O'Connor, M.; Salzman, A. L.; Szabó, C.

1996-01-01

151

Inhibition of melatonin-induced ascorbic acid and LHRH release by a nitric oxide synthase and cyclic GMP inhibitor.  

PubMed

Melatonin (MEL), the principle secretory product of the pineal gland, has been shown to function as an antioxidant and free-radical scavenger. We previously showed that the release of ascorbic acid (AA) and luteinizing hormone releasing hormone (LHRH) from medial basal hypothalamus (MBH) was mediated by nitric oxide (NO) that released cyclic guanosine 3'5'-mono-phosphate (cGMP). Therefore, it was of interest to evaluate the effect of MEL on AA and LHRH release and study the effect of a nitric oxide synthase (NOS) inhibitor, 6-anilino-5,8-quinoline-dione (LY 83583), and a guanylyl cyclase (GC) inhibitor, 1H-[1,2,4] oxadiazolo [4,3-a] quinoxalin-1-one (O.D.Q.), on the release process. Because NO has been shown to activate soluble guanylyl cyclase that elicited an elevation of cGMP in target cells, in the current investigation LY 83583, O.D.Q., or N(G)-monomethyl-l-arginine (NMMA), a competitive inhibitor of NOS, were used to evaluate their effects on MEL-induced AA and LHRH release. Medial basal hypothalami were incubated in 0.5 ml of Krebs-Ringer bicarbonate (KRB) buffer for 1 hr. Subsequently, the tissues were incubated with graded concentrations of MEL (10(-8) to 10(-4) M), MEL + NMMA (3 x 10(-4) M), MEL + LY 83583 (10(-6) M), or MEL + O.D.Q. (10(-5) M) for 1 hr. Ascorbic acid and LHRH released into the medium were measured by high-performance liquid chromatography (HPLC) and radio-immunoassay (RIA), respectively. Melatonin (10(-6) and 10(-5) M) significantly stimulated both AA and LHRH release, but the lower and the highest concentrations were ineffective. A combination of MEL + NMMA completely blocked both AA and LHRH release, supporting a role for NO in the releasing action. Both LY 83583 and O.D.Q. significantly suppressed MEL-induced AA and LHRH release, emphasizing the role of NOS, GC, and cGMP in mediating the action of MEL. The data of these in vitro experiments support a role for MEL in the hypothalamic control of AA and LHRH release. PMID:15229359

Karanth, Sharada; Yu, Wen H; Mastronardi, Claudio A; McCann, Samuel M

2004-07-01

152

Novel benzothiazinones (BTOs) as allosteric modulator or substrate competitive inhibitor of glycogen synthase kinase 3? (GSK-3?) with cellular activity of promoting glucose uptake.  

PubMed

Glycogen synthase kinase 3? (GSK-3?) plays a key role in insulin metabolizing pathway and therefore inhibition of the enzyme might provide an important therapeutic approach for treatment of insulin resistance and type 2 diabetes. Recently, discovery of ATP noncompetitive inhibitors is gaining importance not only due to their generally increased selectivity but also for the potentially subtle modulation of the target. These kinds of compounds include allosteric modulators and substrate competitive inhibitors. Here we reported two benzothiazinone compounds (BTO), named BTO-5h (IC50=8 ?M) and BTO-5s (IC50=10 ?M) as novel allosteric modulator and substrate competitive inhibitor of GSK-3?, respectively. Their different action modes were proved by kinetic experiments. Furthermore, BTO-5s was selected to check the kinases profile and showed little or even no activity to a panel of ten protein kinases at 100 ?M, indicating it has good selectivity. Docking studies were performed to give suggesting binding modes which can well explain their impacts on the enzyme. Moreover, cell experiments displayed both compounds reduced the phosphorylation level of glycogen synthase in an intact cell, and greatly enhanced the glucose uptake in both HpG2 and 3T3-L1 cells. All of these results suggested BTO-5s and BTO-5h maybe have potentially therapeutic value for anti-diabetes. The results also offer a new scaffold for designing and developing selective inhibitors with novel mechanisms of action. PMID:25467150

Zhang, Peng; Li, Shufen; Gao, Yang; Lu, Wenbo; Huang, Ke; Ye, Deyong; Li, Xi; Chu, Yong

2014-12-15

153

An interactional network of genes involved in chitin synthesis in Saccharomyces cerevisiae  

PubMed Central

Background In S. cerevisiae the ?-1,4-linked N-acetylglucosamine polymer, chitin, is synthesized by a family of 3 specialized but interacting chitin synthases encoded by CHS1, CHS2 and CHS3. Chs2p makes chitin in the primary septum, while Chs3p makes chitin in the lateral cell wall and in the bud neck, and can partially compensate for the lack of Chs2p. Chs3p requires a pathway of Bni4p, Chs4p, Chs5p, Chs6p and Chs7p for its localization and activity. Chs1p is thought to have a septum repair function after cell separation. To further explore interactions in the chitin synthase family and to find processes buffering chitin synthesis, we compiled a genetic interaction network of genes showing synthetic interactions with CHS1, CHS3 and genes involved in Chs3p localization and function and made a phenotypic analysis of their mutants. Results Using deletion mutants in CHS1, CHS3, CHS4, CHS5, CHS6, CHS7 and BNI4 in a synthetic genetic array analysis we assembled a network of 316 interactions among 163 genes. The interaction network with CHS3, CHS4, CHS5, CHS6, CHS7 or BNI4 forms a dense neighborhood, with many genes functioning in cell wall assembly or polarized secretion. Chitin levels were altered in 54 of the mutants in individually deleted genes, indicating a functional relationship between them and chitin synthesis. 32 of these mutants triggered the chitin stress response, with elevated chitin levels and a dependence on CHS3. A large fraction of the CHS1-interaction set was distinct from that of the CHS3 network, indicating broad roles for Chs1p in buffering both Chs2p function and more global cell wall robustness. Conclusion Based on their interaction patterns and chitin levels we group interacting mutants into functional categories. Genes interacting with CHS3 are involved in the amelioration of cell wall defects and in septum or bud neck chitin synthesis, and we newly assign a number of genes to these functions. Our genetic analysis of genes not interacting with CHS3 indicate expanded roles for Chs4p, Chs5p and Chs6p in secretory protein trafficking and of Bni4p in bud neck organization. PMID:15715908

Lesage, Guillaume; Shapiro, Jesse; Specht, Charles A; Sdicu, Anne-Marie; Ménard, Patrice; Hussein, Shamiza; Tong, Amy Hin Yan; Boone, Charles; Bussey, Howard

2005-01-01

154

The Discovery of Potentially Selective Human Neuronal Nitric Oxide Synthase (nNOS) Inhibitors: A Combination of Pharmacophore Modelling, CoMFA, Virtual Screening and Molecular Docking Studies  

PubMed Central

Neuronal nitric oxide synthase (nNOS) plays an important role in neurotransmission and smooth muscle relaxation. Selective inhibition of nNOS over its other isozymes is highly desirable for the treatment of neurodegenerative diseases to avoid undesirable effects. In this study, we present a workflow for the identification and prioritization of compounds as potentially selective human nNOS inhibitors. Three-dimensional pharmacophore models were constructed based on a set of known nNOS inhibitors. The pharmacophore models were evaluated by Pareto surface and CoMFA (Comparative Molecular Field Analysis) analyses. The best pharmacophore model, which included 7 pharmacophore features, was used as a search query in the SPECS database (SPECS®, Delft, The Netherlands). The hit compounds were further filtered by scoring and docking. Ten hits were identified as potential selective nNOS inhibitors. PMID:24830557

Xu, Guanhong; Chen, Yue; Shen, Kun; Wang, Xiuzhen; Li, Fei; He, Yan

2014-01-01

155

Protective Role of Selective Nitric Oxide Synthase Inhibitor for Treatment of Decompensated Hemorrhagic Shock in Normotensive and Hypertensive Rats  

PubMed Central

Introduction: Different vasoactive factors can modulate cardiovascular adaptation to hemorrhagic shock including Nitric Oxide (NO). In this study we investigated the effect of the NO synthase inhibitor for treatment of decompensated hemorrhagic shock in normotensive and hypertensive rats. Methods: Twenty-four male Wistar rats were divided into two groups: The normotensive and hypertensive groups. Hypertension was induced by the DOCA-Salt method for eight weeks. Then, the animals were given hemorrhagic shock by continuously withdrawing blood until the mean arterial pressure (MAP) reached to 40 mmHg. The animals were maintained in the shock state for 120 minutes. Subsequently, they were randomly assigned to L-NAME-treated and non-treated groups and monitored for 60 minutes. The survival time was recorded. Blood samples were taken before and after the shock and 60 minutes after L-NAME administration. Results: Infusion of L-NAME caused a significant increase in MAP in normotensive animals, however, slightly increased MAP in hypertensive animals. The heart rate did not significantly alter. Hemorrhage caused a marked increase in serum nitrite levels in both groups (P<0.05). L-NAME treatment significantly reduced the serum nitrite concentration in the normotensive group (P<0.05), without any change in the hypertensive group. All animals who received L-NAME treatment survived at the end of experiment. Fifty percent of the hypertensive animals died four hours after the experiment. The 72-hour survival rate was similar in the L-NAME treated groups. Conclusion: L-NAME infusion during decompensated hemorrhagic shock plays a protective role in the improvement of hemodynamic responses and short-term survival rate in normotensive animals. PMID:22355477

Khazaei, Majid; Barmaki, Babak; Nasimi, Ali

2012-01-01

156

Protective effects of poly (ADP-ribose) synthase inhibitors on digoxin-induced cardiotoxicity in guinea-pig isolated hearts.  

PubMed

Reactive oxygen species, generated and released during digoxin-induced cardiotoxicity, can produce an activation of poly (ADP-ribose) synthase (PARS). Our objective was to examine the effects of PARS inhibitors, 3-aminobenzamide (3-AB ) and nicotinamide, on digoxin-induced arrhythmias in guinea-pig isolated hearts. 3-AB (0.1-0.3 mM) and nicotinamide (0.3 mM) were added to the perfusion solution starting 10 min before digoxin infusion (8 microg x ml (-1)min (-1)reaching the heart) and maintained throughout the experiments. Electrocardiograms and coronary perfusion pressure were recorded continuously, and digoxin-induced arrhythmias were determined. Nicotinamide markedly inhibited ventricular tachycardia (VT) incidence (from 100%, n= 7, to 29%, n= 7), and abolished ventricular fibrillation (VF) incidence. 3-AB (0.1 mM, n= 9) significantly decreased VT incidence from 100% ( n= 7) to 22% ( n= 9) and VF incidence from 86% ( n= 7) to 11% ( n= 9). Both nicotinamide and 3-AB (0.1 mM) markedly decreased number of ventricular ectopic beats (VEBs) and arrhythmia score. 3-AB at 0.3 mM ( n= 8) appeared to decrease the VT (to 63%) and VF incidence (to 38%), but these reductions did not reach statistically significance levels. Moreover, 3-AB at high concentration (0.3 mM) did not significantly modify the number of VEBs and arrhythmia score. There were no significant changes in coronary perfusion pressure, heart rate or pressure rate index measured at certain time points throughout the experiment in all groups. Our results suggest that PARS activation plays a role in the digitalis-induced cardiotoxicity in guinea-pig isolated hearts. PMID:11884214

Demiryürek, A Tuncay; Yildiz, Gülüzar; E?iyok, Sibel; Altu?, Sedat

2002-03-01

157

CESA TRAFFICKING INHIBITOR Inhibits Cellulose Deposition and Interferes with the Trafficking of Cellulose Synthase Complexes and Their Associated Proteins KORRIGAN1 and POM2/CELLULOSE SYNTHASE INTERACTIVE PROTEIN1.  

PubMed

Cellulose synthase complexes (CSCs) at the plasma membrane (PM) are aligned with cortical microtubules (MTs) and direct the biosynthesis of cellulose. The mechanism of the interaction between CSCs and MTs, and the cellular determinants that control the delivery of CSCs at the PM, are not yet well understood. We identified a unique small molecule, CESA TRAFFICKING INHIBITOR (CESTRIN), which reduces cellulose content and alters the anisotropic growth of Arabidopsis (Arabidopsis thaliana) hypocotyls. We monitored the distribution and mobility of fluorescently labeled cellulose synthases (CESAs) in live Arabidopsis cells under chemical exposure to characterize their subcellular effects. CESTRIN reduces the velocity of PM CSCs and causes their accumulation in the cell cortex. The CSC-associated proteins KORRIGAN1 (KOR1) and POM2/CELLULOSE SYNTHASE INTERACTIVE PROTEIN1 (CSI1) were differentially affected by CESTRIN treatment, indicating different forms of association with the PM CSCs. KOR1 accumulated in bodies similar to CESA; however, POM2/CSI1 dissociated into the cytoplasm. In addition, MT stability was altered without direct inhibition of MT polymerization, suggesting a feedback mechanism caused by cellulose interference. The selectivity of CESTRIN was assessed using a variety of subcellular markers for which no morphological effect was observed. The association of CESAs with vesicles decorated by the trans-Golgi network-localized protein SYNTAXIN OF PLANTS61 (SYP61) was increased under CESTRIN treatment, implicating SYP61 compartments in CESA trafficking. The properties of CESTRIN compared with known CESA inhibitors afford unique avenues to study and understand the mechanism under which PM-associated CSCs are maintained and interact with MTs and to dissect their trafficking routes in etiolated hypocotyls. PMID:25535279

Worden, Natasha; Wilkop, Thomas E; Esteve, Victor Esteva; Jeannotte, Richard; Lathe, Rahul; Vernhettes, Samantha; Weimer, Bart; Hicks, Glenn; Alonso, Jose; Labavitch, John; Persson, Staffan; Ehrhardt, David; Drakakaki, Georgia

2015-02-01

158

MICROBIOLOGY: Chitin, Cholera, and Competence  

NSDL National Science Digital Library

Access to the article is free, however registration and sign-in are required. Vibrio cholerae, a human pathogen, inhabits aquatic environments and is often associated with chitin-containing organisms. In their Perspective, Bartlett and Azam discuss the findings of Meibom et al. in the same issue of chitin-mediated natural DNA transformation in V. cholerae. This finding opens a new window for understanding conditions that influence the evolution of this bacterium in aquatic habitats.

Douglas H. Bartlett (cripps Institution of Oceanography, University of California;Marine Biology Research Division); Farooq Azam (cripps Institution of Oceanography, University of California;Marine Biology Research Division)

2005-12-16

159

L-Vinylglycine is an alternative substrate as well as a mechanism-based inhibitor of 1-aminocyclopropane-1-carboxylate synthase.  

PubMed

L-Vinylglycine (L-VG) has been shown to be a mechanism-based inhibitor of 1-aminocyclopropane-1-carboxylate (ACC) synthase [Satoh, S., and Yang, S. F. (1989) Plant Physiol. 91, 1036-1039] as well as of other pyridoxal phosphate-dependent enzymes. This report demonstrates that L-VG is primarily an alternative substrate for the enzyme. The L-VG deaminase activity of ACC synthase yields the products alpha-ketobutyrate and ammonia with a k(cat) value of 1.8 s(-1) and a K(m) value of 1.4 mM. The k(cat)/K(m) of 1300 M(-1) s(-1) is 0.17% that of the diffusion-controlled reaction with the preferred substrate, S-adenosyl-L-methionine. The enzyme-L-VG complex partitions to products 500 times for every inactivation event. The catalytic mechanism proceeds through a spectrophotometrically detected quinonoid with lambda(max) of 530 nm, which must rearrange to a 2-aminocrotonate aldimine to yield final products. Alternative mechanisms for the inactivation reaction are presented, and the observed kinetics for the full reaction course are satisfactorily modeled by kinetic simulation. The inactive enzyme is an aldimine with lambda(max) of 432 nm. It is resistant to NaBH(3)CN but is reduced by NaBH(4). ACC synthase is now expressed in Pichia pastoris with an improved yield of 10 mg/L. PMID:10704193

Feng, L; Kirsch, J F

2000-03-14

160

Chs7p, a New Protein Involved in the Control of Protein Export from the Endoplasmic Reticulum that Is Specifically Engaged in the Regulation of Chitin Synthesis in Saccharomyces cerevisiae  

Microsoft Academic Search

The Saccharomyces cerevisiae CHS7 gene encodes an integral membrane protein located in the ER which is directly involved in chitin synthesis through the regulation of chitin synthase III (CSIII) activity. In the absence of CHS7 product, Chs3p, but not other secreted proteins, is retained in the ER, lead- ing to a severe defect in CSIII activity and conse- quently, to

Jose A. Trilla; Angel Durán; Cesar Roncero

1999-01-01

161

Multivariate SAR/QSAR of 3-aryl-4-hydroxyquinolin-2(1H)-one derivatives as type I fatty acid synthase (FAS) inhibitors.  

PubMed

Two multivariate studies, a PCA-SAR and a PLS-QSAR, of 3-aryl-4-hydroxyquinolin-2(1H)-one derivatives described as type I fatty acid synthase (FAS) inhibitors, are presented in this work. The variable selection was performed with the Fisher's weight and Ordered Predictors Selection (OPS) algorithm, respectively. In the PCA, a separation between active and inactive compounds was obtained by six descriptors (topological and geometrical). The PLS model presented five descriptors and two Latent Variables. Leave-N-out cross validation and y-randomization test showed that the model presented robustness and no chance correlation, respectively, and the descriptors indicated that the FAS inhibition depends on electronic distribution of the investigated compounds. The model obtained in this study may provide a guidance for proposition of new FAS inhibitors. PMID:20965618

de Melo, Eduardo Borges

2010-12-01

162

Interactions between inducible isoforms of nitric oxide synthase and cyclo-oxygenase in vivo: investigations using the selective inhibitors, 1400W and celecoxib  

PubMed Central

Exposure of tissues to endotoxin (LPS) and/or cytokines leads to the induction of both inducible nitric oxide synthase (iNOS) and cyclo-oxygenase-2 (COX-2). It has previously been reported that there is `cross-talk' between these two systems. However, such previous studies have been limited by the availability of highly selective inhibitors. Here we have investigated the interactions between iNOS and COX-2 in vivo using 1400W, an iNOS-selective inhibitor, and celecoxib, a COX-2-selective inhibitor.Infusion of LPS to rats for 6?h caused a time-dependent increase in the plasma concentrations of 6 keto-prostaglandin F1? (6 keto-PGF1?) and nitrite/nitrate (NO2/NO3), consistent with the induction of iNOS and COX-2. Bolus injection of arachidonic acid (AA) at t=6?h resulted in a further increase of circulating levels of 6 keto-PGF1? in LPS-treated animals.Treatment of rats with 1400W or the non-selective NOS inhibitor NG-monomethyl-L-arginine (L-NMMA) inhibited the increase in plasma NO2/NO3 but were both without effect on the plasma concentration of 6 keto-PGF1? before or after AA.Treatment with the non-steroidal anti-inflammatory drugs (NSAIDs), A771726 or diclofenac, or with celecoxib significantly reduced the increase in circulating 6 keto-PGF1? caused by LPS, and the large increase in 6 keto-PGF1? following injection of AA. None of the COX inhibitors affected the increase in plasma NO2/NO3. Dexamethasone, however, significantly inhibited both the increase in 6 keto-PGF1? and the increase in NO2/NO3.In conclusion, the use of selective inhibitors does not support the concept of cross talk in vivo between iNOS and COX-2. PMID:9786506

Hamilton, Lorna C; Warner, Timothy D

1998-01-01

163

1,2-Dithiole-3-Ones as Potent Inhibitors of the Bacterial 3-Ketoacyl Acyl Carrier Protein Synthase III (FabH)  

PubMed Central

The enzyme FabH catalyzes the initial step of fatty acid biosynthesis via a type II dissociated fatty acid synthase. The pivotal role of this essential enzyme, combined with its unique structural features and ubiquitous occurrence in bacteria, has made it an attractive new target for the development of antibacterial and antiparasitic compounds. We have searched the National Cancer Institute database for compounds bearing structural similarities to thiolactomycin, a natural product which exhibits a weak activity against FabH. This search has yielded several substituted 1,2-dithiole-3-ones that are potent inhibitors of FabH from both Escherichia coli (ecFabH) and Staphylococcus aureus (saFabH). The most potent inhibitor was 4,5-dichloro-1,2-dithiole-3-one, which had 50% inhibitory concentration (IC50) values of 2 ?M (ecFabH) and 0.16 ?M (saFabH). The corresponding 3-thione analog exhibited comparable activities. Analogs in which the 4-chloro substituent was replaced with a phenyl group were also potent inhibitors, albeit somewhat less effectively (IC50 values of 5.7 and 0.98 ?M for ecFabH and saFabH, respectively). All of the 5-chlorinated inhibitors were most effective when they were preincubated with FabH in the absence of substrates. The resulting enzyme-inhibitor complex did not readily regain activity after excess inhibitor was removed, suggesting that a slow dissociation occurs. In stark contrast, a series of inhibitors in which the 5-chloro substituent was replaced with the isosteric and isoelectronic trifluoromethyl group were poorer inhibitors (IC50 values typically ranging from 25 to >100 ?M for both ecFabH and saFabH), did not require a preincubation period for maximal activity, and generated an enzyme-inhibitor complex which readily dissociated. Possible modes of binding of 5-chloro-1,2-dithiole-3-ones and 5-chloro-1,2-dithiole-3-thiones with FabH which account for the role of the 5-chloro substituent were considered. PMID:15273125

He, Xin; Reeve, Anne McElwee; Desai, Umesh R.; Kellogg, Glen E.; Reynolds, Kevin A.

2004-01-01

164

Chitin deposition on the embryonic cuticle of Rhodnius prolixus: the reduction of CHS transcripts by CHS-dsRNA injection in females affects chitin deposition and eclosion of the first instar nymph.  

PubMed

In a previous study, we found that the embryonic cuticle of Rhodnius prolixus is a chitin-based structure that helps the first instar nymph to hatch from the chorion. Here, we investigated how the reduction of transcripts induced by CHS dsRNA injection affects R. prolixus embryogenesis and eclosion. Deposition of chitin in the embryonic cuticle begins later at embryogenesis, around day 8, and ends approximately at day 15, when the insects are ready for eclosion. In R. prolixus, chitin deposition follows pari passu with the synthesis of the chitin synthase mRNA, indicating a regulation at the transcriptional level. The reduction of the chitin synthase gene transcripts by the injection of CHS dRNA prevented chitin deposition during embryonic cuticle formation, being lethal to hatching nymphs, which end up dying while stuck in the chorionic border trying to leave the chorion. The successful eclosion rates were reduced by 60% in animals treated with CHS dsRNA when compared to animals injected with a control (dsRNA no related gene or water). We found that the harmful effects on oviposition and eclosion are possibly due to changes in the structure of the embryonic cuticle, as observed by directly comparing the morphology of control and chitin-deficient embryonic cuticles under the transmission electron microscope. The lack of chitin and changes in its morphological characteristics appears to alter the embryonic cuticle physiology and functionality. Additionally, we observed that the effects of CHS dRNA treatment on R. prolixus females lasted up to 3 egg-laying cycles (?100 days), pointing to R. prolixus as a useful model for developmental studies. PMID:24412274

Souza-Ferreira, Paula S; Mansur, Juliana F; Berni, Matheus; Moreira, Monica F; dos Santos, Roberto Eizemberg; Araújo, Helena M Marcolla; de Souza, Wanderley; Ramos, Isabela B; Masuda, Hatisaburo

2014-08-01

165

A novel role of andrographolide, an NF-kappa B inhibitor, on inhibition of platelet activation: the pivotal mechanisms of endothelial nitric oxide synthase/cyclic GMP.  

PubMed

Andrographolide is a novel NF-?B inhibitor from the leaves of Andrographis paniculata. Platelet activation is relevant to a variety of thrombotic diseases. However, no data are available concerning the effects of andrographolide in platelet activation. The aim of this study was to examine the mechanisms of andrographolide in preventing platelet activation. Andrographolide (25-75 ??) exhibited a more potent activity of inhibiting platelet aggregation stimulated by collagen. Andrographolide inhibited collagen-stimulated platelet activation accompanied by relative Ca(2+) mobilization; thromboxane A(2) formation; and phospholipase C (PLC)?2, protein kinase C (PKC), mitogen-activated protein kinase (MAPK), and Akt phosphorylation. Andrographolide markedly increased cyclic GMP, but not cyclic AMP levels. Andrographolide also stimulated endothelial nitric oxide synthase (eNOS) expression, NO release, and vasodilator-stimulated phosphoprotein (VASP) phosphorylation. ODQ, an inhibitor of guanylate cyclase, markedly reversed the andrographolide-mediated inhibitory effects on platelet aggregation, p38 MAPK and Akt phosphorylation, and the andrographolide-mediated stimulatory effect on VASP phosphorylation. Furthermore, a PI3 kinase inhibitor (LY294002) but not a PKC inhibitor (Ro318220) significantly diminished p38 MAPK phosphorylation; nevertheless, a p38 MAPK inhibitor (SB203580) and LY294002 diminished PKC activity stimulated by collagen. Andrographolide also reduced collagen-triggered hydroxyl radical (OH([Symbol: see text])) formation. In vivo studies revealed that andrographolide (22 and 55 ?g/kg) is effective in reducing the mortality of ADP-induced acute pulmonary thromboembolism and significantly prolonged platelet plug formation in mice. This study demonstrates for the first time that andrographolide possesses a novel role of antiplatelet activity, which may involve the activation of the eNOS-NO/cyclic GMP pathway, resulting in the inhibition of the PI3 kinase/Akt-p38 MAPK and PLC?2-PKC cascades, thereby leading to inhibition of platelet aggregation. PMID:21822619

Lu, Wan-Jung; Lee, Jie-Jen; Chou, Duen-Suey; Jayakumar, Thanasekaran; Fong, Tsorng-Han; Hsiao, George; Sheu, Joen-Rong

2011-12-01

166

Activation of ?-catenin by inhibitors of glycogen synthase kinase-3 ameliorates cisplatin-induced cytotoxicity and pro-inflammatory cytokine expression in HEI-OC1 cells.  

PubMed

Cisplatin is used in the treatment of a wide variety of solid tumors, but its use is limited by its serious adverse effects, including ototoxicity. Glycogen synthase kinase-3 (GSK-3) is a ubiquitously expressed serine/threonine kinase that regulates a variety of cellular functions by phosphorylating its substrates. However, the otoprotective effect of GSK-3 inhibitors is poorly understood. Here, we investigated whether GSK-3 is involved in cisplatin-induced ototoxicity in HEI-OC1 cells and organs of Corti (OCs). GSK-3 inhibitors suppressed cisplatin-induced apoptosis determined by decreased p53 activity, and also decreased expression of PARP and p53 target genes such as p21 and PUMA. The effect of GSK-3 inhibitors was mediated by markedly increased nuclear ?-catenin that in turn blocked nuclear translocation of NF-?B. siRNA-mediated ?-catenin knockdown markedly increased the expression of NF-?B target genes, such as TNF-? and IL-6. Our data suggest that the GSK-3/?-catenin pathway may play a central role in cisplatin-mediated cytotoxicity in HEI-OC1 cells and hair cells of OCs in vitro. PMID:24560772

Kim, Se-Jin; Lim, Jae-Young; Lee, Joon No; Choe, Seong-Kyu; Kim, Yong-Il; Song, Seung Ryel; Cho, Meyoung; So, Hong-Seob; Park, Raekil

2014-06-01

167

In-silico docking based design and synthesis of [1H,3H] imidazo[4,5-b] pyridines as lumazine synthase inhibitors for their effective antimicrobial activity  

PubMed Central

Purpose: The imidazopyridine moiety is important pharmacophore that has proven to be useful for a number of biologically relevant targets, also reported to display antibacterial, antifungal, antiviral properties. Riboflavin biosynthesis involving catalytic step of Lumazine synthase is absent in animals and human, but present in microorganism, one of marked advantage of this study. Still, this path is not exploited as antiinfective target. Here, we proposed different interactions between [1H,3H] imidazo[4,5-b] pyridine test ligands and target protein Lumazine synthase (protein Data Bank 2C92), one-step synthesis of title compounds and further evaluation of them for in vitro antimicrobial activity. Materials and Methods: Active pocket of the target protein involved in the interaction with the test ligands molecules was found using Biopredicta tools in VLifeMDS 4.3 Suite. In-silico docking suggests H-bonding, hydrophobic interaction, charge interaction, aromatic interaction, and Vanderwaal forces responsible for stabilizing enzyme-inhibitor complex. Disc diffusion assay method was used for in vitro antimicrobial screening. Results and Discussion: Investigation of possible interaction between test ligands and target lumazine synthase of Mycobacterium tuberculosis suggested 1i and 2f as best fit candidates showing hydrogen bonding, hydrophobic, aromatic and Vanderwaal's forces. Among all derivatives 1g, 1j, 1k, 1l, 2a, 2c, 2d, 2e, 2h, and 2j exhibited potent activities against bacteria and fungi compared to the standard Ciprofloxacin and Fluconazole, respectively. The superiority of 1H imidazo [4,5-b] pyridine compounds having R’ = Cl >No2 > NH2 at the phenyl/aliphatic moiety resident on the imidazopyridine, whereas leading 3H imidazo[4,5-b] pyridine compounds containing R/Ar = Cl > No2 > NH2> OCH3 substituents on the 2nd position of imidazole. PMID:25400412

Harer, Sunil L.; Bhatia, Manish S.

2014-01-01

168

Effects of the dual TP receptor antagonist and thromboxane synthase inhibitor EV-077 on human endothelial and vascular smooth muscle cells.  

PubMed

The prothrombotic mediator thromboxane A2 is derived from arachidonic acid metabolism through the cyclooxygenase and thromboxane synthase pathways, and transduces its effect through the thromboxane prostanoid (TP) receptor. The aim of this study was to determine the effect of the TP receptor antagonist and thromboxane synthase inhibitor EV-077 on inflammatory markers in human umbilical vein endothelial cells and on human coronary artery smooth muscle cell proliferation. To this end, mRNA levels of different proinflammatory mediators were studied by real time quantitative PCR, supernatants were analyzed by enzyme immune assay, and cell proliferation was assessed using WST-1. EV-077 significantly decreased mRNA levels of ICAM-1 and PTX3 after TNF? incubation, whereas concentrations of 6-keto PGF1? in supernatants of endothelial cells incubated with TNF? were significantly increased after EV-077 treatment. Although U46619 did not alter coronary artery smooth muscle cell proliferation, this thromboxane mimetic enhanced the proliferation induced by serum, insulin and growth factors, which was significantly inhibited by EV-077. In conclusion, EV-077 inhibited TNF?-induced endothelial inflammation and reduced the enhancement of smooth muscle cell proliferation induced by a thromboxane mimetic, supporting that the thromboxane pathway may be associated with early atherosclerosis in terms of endothelial dysfunction and vascular hypertrophy. PMID:24161392

Petri, Marcelo H; Tellier, Céline; Michiels, Carine; Ellertsen, Ingvill; Dogné, Jean-Michel; Bäck, Magnus

2013-11-15

169

Effects of the dual TP receptor antagonist and thromboxane synthase inhibitor EV-077 on human endothelial and vascular smooth muscle cells  

SciTech Connect

Highlights: •EV-077 reduced TNF-? induced inflammation in endothelial cells. •The thromboxane mimetic U69915 enhanced vascular smooth muscle cell proliferation. •EV-077 inhibited smooth muscle cell proliferation. -- Abstract: The prothrombotic mediator thromboxane A{sub 2} is derived from arachidonic acid metabolism through the cyclooxygenase and thromboxane synthase pathways, and transduces its effect through the thromboxane prostanoid (TP) receptor. The aim of this study was to determine the effect of the TP receptor antagonist and thromboxane synthase inhibitor EV-077 on inflammatory markers in human umbilical vein endothelial cells and on human coronary artery smooth muscle cell proliferation. To this end, mRNA levels of different proinflammatory mediators were studied by real time quantitative PCR, supernatants were analyzed by enzyme immune assay, and cell proliferation was assessed using WST-1. EV-077 significantly decreased mRNA levels of ICAM-1 and PTX3 after TNF? incubation, whereas concentrations of 6-keto PGF1? in supernatants of endothelial cells incubated with TNF? were significantly increased after EV-077 treatment. Although U46619 did not alter coronary artery smooth muscle cell proliferation, this thromboxane mimetic enhanced the proliferation induced by serum, insulin and growth factors, which was significantly inhibited by EV-077. In conclusion, EV-077 inhibited TNF?-induced endothelial inflammation and reduced the enhancement of smooth muscle cell proliferation induced by a thromboxane mimetic, supporting that the thromboxane pathway may be associated with early atherosclerosis in terms of endothelial dysfunction and vascular hypertrophy.

Petri, Marcelo H. [Department of Medicine, Karolinska Institutet and Center for Molecular Medicine, Karolinska University Hospital, Stockholm (Sweden)] [Department of Medicine, Karolinska Institutet and Center for Molecular Medicine, Karolinska University Hospital, Stockholm (Sweden); Tellier, Céline; Michiels, Carine [NARILIS, URBC, University of Namur, Namur (Belgium)] [NARILIS, URBC, University of Namur, Namur (Belgium); Ellertsen, Ingvill [Department of Medicine, Karolinska Institutet and Center for Molecular Medicine, Karolinska University Hospital, Stockholm (Sweden)] [Department of Medicine, Karolinska Institutet and Center for Molecular Medicine, Karolinska University Hospital, Stockholm (Sweden); Dogné, Jean-Michel [Department of Pharmacy, Namur Thrombosis and Hemostasis Center, University of Namur, Namur (Belgium)] [Department of Pharmacy, Namur Thrombosis and Hemostasis Center, University of Namur, Namur (Belgium); Bäck, Magnus, E-mail: Magnus.Back@ki.se [Department of Medicine, Karolinska Institutet and Center for Molecular Medicine, Karolinska University Hospital, Stockholm (Sweden)] [Department of Medicine, Karolinska Institutet and Center for Molecular Medicine, Karolinska University Hospital, Stockholm (Sweden)

2013-11-15

170

Comparative evaluation of the efficacy of the cyclooxygenase pathway inhibitor and nitric oxide synthase inhibitor in the reduction of alveolar bone loss in ligature induced periodontitis in rats: An experimental study  

PubMed Central

Background: Alveolar bone loss is the most striking feature of periodontal disease. The aim of this study was to investigate the effect of a cyclooxygenase (COX) pathway inhibitor and nitric oxide synthase (NOS) inhibitor in the reduction of alveolar bone loss in an experimental periodontal disease (EPD) model. Materials and Methods: The study was conducted on 60 Wistar rats divided into three groups of 20 rats each and then subjected to a ligature placement around the left maxillary second molars. Group 1 rats were treated with COX inhibitor (diclofenac sodium 10 mg/kg/d), group 2 with NOS inhibitor (aminoguanidine hydrochloride 10 mg/kg/d) and group 3 served as controls, receiving only saline, intraperitoneally 1h before EPD induction and daily until the sacrifice on the 11th day. Leukogram was performed before ligation, at 6 h and at the first, seventh and 11th days after EPD induction. After sacrifice, all the excised maxillae were subjected to morphometric and histometric analysis to measure the alveolar bone loss. Histopathological analysis was carried out to estimate cell influx, alveolar bone and cementum integrity. Results: Induction of experimental periodontitis in the rat model produced pronounced leucocytosis, which was significantly reduced by the administration of diclofenac sodium and aminoguanidine on the 11th day. In morphometric and histometric examinations, both the test drugs significantly (P < 0.05) inhibited the alveolar bone loss as compared with the control group. Conclusion: Both COX inhibitor and NOS inhibitor are equally effective in inhibiting the inflammatory bone resorption in an experimental periodontitis model. PMID:24744546

Jagadish, Rekha; Mehta, Dhoom Singh

2014-01-01

171

Chitin-Like Molecules Associate with Cryptococcus neoformans Glucuronoxylomannan To Form a Glycan Complex with Previously Unknown Properties  

PubMed Central

In prior studies, we demonstrated that glucuronoxylomannan (GXM), the major capsular polysaccharide of the fungal pathogen Cryptococcus neoformans, interacts with chitin oligomers at the cell wall-capsule interface. The structural determinants regulating these carbohydrate-carbohydrate interactions, as well as the functions of these structures, have remained unknown. In this study, we demonstrate that glycan complexes composed of chitooligomers and GXM are formed during fungal growth and macrophage infection by C. neoformans. To investigate the required determinants for the assembly of chitin-GXM complexes, we developed a quantitative scanning electron microscopy-based method using different polysaccharide samples as inhibitors of the interaction of chitin with GXM. This assay revealed that chitin-GXM association involves noncovalent bonds and large GXM fibers and depends on the N-acetyl amino group of chitin. Carboxyl and O-acetyl groups of GXM are not required for polysaccharide-polysaccharide interactions. Glycan complex structures composed of cryptococcal GXM and chitin-derived oligomers were tested for their ability to induce pulmonary cytokines in mice. They were significantly more efficient than either GXM or chitin oligomers alone in inducing the production of lung interleukin 10 (IL-10), IL-17, and tumor necrosis factor alpha (TNF-?). These results indicate that association of chitin-derived structures with GXM through their N-acetyl amino groups generates glycan complexes with previously unknown properties. PMID:22562469

Ramos, Caroline L.; Fonseca, Fernanda L.; Rodrigues, Jessica; Guimarães, Allan J.; Cinelli, Leonardo P.; Miranda, Kildare; Nimrichter, Leonardo; Casadevall, Arturo; Travassos, Luiz R.

2012-01-01

172

Kinase Inhibitor Screening Identifies Cyclin-Dependent Kinases and Glycogen Synthase Kinase 3 as Potential Modulators of TDP-43 Cytosolic Accumulation during Cell Stress  

PubMed Central

Abnormal processing of TAR DNA binding protein 43 (TDP-43) has been identified as a major factor in neuronal degeneration during amyotrophic lateral sclerosis (ALS) or frontotemporal lobar degeneration (FTLD). It is unclear how changes to TDP-43, including nuclear to cytosolic translocation and subsequent accumulation, are controlled in these diseases. TDP-43 is a member of the heterogeneous ribonucleoprotein (hnRNP) RNA binding protein family and is known to associate with cytosolic RNA stress granule proteins in ALS and FTLD. hnRNP trafficking and accumulation is controlled by the action of specific kinases including members of the mitogen-activated protein kinase (MAPK) pathway. However, little is known about how kinase pathways control TDP-43 movement and accumulation. In this study, we used an in vitro model of TDP-43-positve stress granule formation to screen for the effect of kinase inhibitors on TDP-43 accumulation. We found that while a number of kinase inhibitors, particularly of the MAPK pathways modulated both TDP-43 and the global stress granule marker, human antigen R (HuR), multiple inhibitors were more specific to TDP-43 accumulation, including inhibitors of cyclin-dependent kinases (CDKs) and glycogen synthase kinase 3 (GSK3). Close correlation was observed between effects of these inhibitors on TDP-43, hnRNP K and TIAR, but often with different effects on HuR accumulation. This may indicate a potential interaction between TDP-43, hnRNP K and TIAR. CDK inhibitors were also found to reverse pre-formed TDP-43-positive stress granules and both CDK and GSK3 inhibitors abrogated the accumulation of C-terminal TDP-43 (219–414) in transfected cells. Further studies are required to confirm the specific kinases involved and whether their action is through phosphorylation of the TDP-43 binding partner hnRNP K. This knowledge provides a valuable insight into the mechanisms controlling abnormal cytoplasmic TDP-43 accumulation and may herald new opportunities for kinase modulation-based therapeutic intervention in ALS and FTLD. PMID:23840699

Moujalled, Diane; James, Janine L.; Parker, Sarah J.; Lidgerwood, Grace E.; Duncan, Clare; Meyerowitz, Jodi; Nonaka, Takashi; Hasegawa, Masato; Kanninen, Katja M.; Grubman, Alexandra; Liddell, Jeffrey R.; Crouch, Peter J.; White, Anthony R.

2013-01-01

173

Embryonic desiccation resistance in Aedes aegypti: presumptive role of the chitinized Serosal Cuticle  

PubMed Central

Background One of the major problems concerning dengue transmission is that embryos of its main vector, the mosquito Aedes aegypti, resist desiccation, surviving several months under dry conditions. The serosal cuticle (SC) contributes to mosquito egg desiccation resistance, but the kinetics of SC secretion during embryogenesis is unknown. It has been argued that mosquito SC contains chitin as one of its components, however conclusive evidence is still missing. Results We observed an abrupt acquisition of desiccation resistance during Ae. aegypti embryogenesis associated with serosal cuticle secretion, occurring at complete germ band extension, between 11 and 13 hours after egglaying. After SC formation embryos are viable on dry for at least several days. The presence of chitin as one of the SC constituents was confirmed through Calcofluor and WGA labeling and chitin quantitation. The Ae. aegypti Chitin Synthase A gene (AaCHS1) possesses two alternatively spliced variants, AaCHS1a and AaCHS1b, differentially expressed during Ae. aegypti embryonic development. It was verified that at the moment of serosal cuticle formation, AaCHS1a is the sole variant specifically expressed. Conclusion In addition to the peritrophic matrix and exoskeleton, these findings confirm chitin is also present in the mosquito serosal cuticle. They also point to the role of the chitinized SC in the desiccation resistance of Ae. aegypti eggs. AaCHS1a expression would be responsible for SC chitin synthesis. With this embryological approach we expect to shed new light regarding this important physiological process related to the Ae. aegypti life cycle. PMID:18789161

Rezende, Gustavo Lazzaro; Martins, Ademir Jesus; Gentile, Carla; Farnesi, Luana Cristina; Pelajo-Machado, Marcelo; Peixoto, Alexandre Afrânio; Valle, Denise

2008-01-01

174

In vitro activities of ER-119884 and E5700, two potent squalene synthase inhibitors, against Leishmania amazonensis: antiproliferative, biochemical, and ultrastructural effects.  

PubMed

ER-119884 and E5700, novel arylquinuclidine derivatives developed as cholesterol-lowering agents, were potent in vitro growth inhibitors of both proliferative stages of Leishmania amazonensis, the main causative agent of cutaneous leishmaniasis in South America, with the 50% inhibitory concentrations (IC(50)s) being in the low-nanomolar to subnanomolar range. The compounds were very potent noncompetitive inhibitors of native L. amazonensis squalene synthase (SQS), with inhibition constants also being in the nanomolar to subnanomolar range. Growth inhibition was strictly associated with the depletion of the parasite's main endogenous sterols and the concomitant accumulation of exogenous cholesterol. Using electron microscopy, we identified the intracellular structures affected by the compounds. A large number of lipid inclusions displaying different shapes and electron densities were observed after treatment with both SQS inhibitors, and these inclusions were associated with an intense disorganization of the membrane that surrounds the cell body and flagellum, as well as the endoplasmic reticulum and the Golgi complex. Cells treated with ER-119884 but not those treated with E5700 had an altered cytoskeleton organization due to an abnormal distribution of tubulin, and many were arrested at cytokinesis. A prominent contractile vacuole and a phenotype typical of programmed cell death were frequently found in drug-treated cells. The selectivity of the drugs was demonstrated with the JC-1 mitochondrial fluorescent label and by trypan blue exclusion tests with macrophages, which showed that the IC(50)s against the host cells were 4 to 5 orders of magnitude greater that those against the intracellular parasites. Taken together, our results show that ER-119884 and E5700 are unusually potent and selective inhibitors of the growth of Leishmania amazonensis, probably because of their inhibitory effects on de novo sterol biosynthesis at the level of SQS, but some of our observations indicate that ER-119884 may also interfere with other cellular processes. PMID:18765694

Fernandes Rodrigues, Juliany Cola; Concepcion, Juan Luis; Rodrigues, Carlos; Caldera, Aura; Urbina, Julio A; de Souza, Wanderley

2008-11-01

175

Distinction of microsomal prostaglandin E synthase-1 (mPGES-1) inhibition from cyclooxygenase-2 inhibition in cells using a novel, selective mPGES-1 inhibitor.  

PubMed

Inflammation-induced microsomal prostaglandin E synthase-1 (mPGES-1) is the terminal enzyme that synthesizes prostaglandin E(2) (PGE(2)) downstream of cyclooxygenase-2 (COX-2). The efficacy of nonsteroidal anti-inflammatory drugs and COX-2 inhibitors in the treatment of the signs and symptoms of osteoarthritis, rheumatoid arthritis and inflammatory pain, largely attributed to the inhibition of PGE(2) synthesis, provides a rationale for exploring mPGES-1 inhibition as a potential novel therapy for these diseases. Toward this aim, we identified PF-9184 as a novel mPGES-1 inhibitor. PF-9184 potently inhibited recombinant human (rh) mPGES-1 (IC(50)=16.5+/-3.8nM), and had no effect against rhCOX-1 and rhCOX-2 (>6500-fold selectivity). In inflammation and clinically relevant biological systems, mPGES-1 expression, like COX-2 expression was induced in cell context- and time-dependent manner, consistent with the kinetics of PGE(2) synthesis. In rationally designed cell systems ideal for determining direct effects of the inhibitors on mPGES-1 function, but not its expression, PF-9184 inhibited PGE(2) synthesis (IC(50) in the range of 0.5-5 microM in serum-free cell and human whole blood cultures, respectively) while sparing the synthesis of 6-keto-PGF(1alpha) (PGF(1alpha)) and PGF(2alpha). In contrast, as expected, the selective COX-2 inhibitor, SC-236, inhibited PGE(2), PGF(1alpha) and PGF(2alpha) synthesis. This profile of mPGES-1 inhibition, distinct from COX-2 inhibition in cells, validates mPGES-1 as an attractive target for therapeutic intervention. PMID:20067770

Mbalaviele, Gabriel; Pauley, Adele M; Shaffer, Alexander F; Zweifel, Ben S; Mathialagan, Sumathy; Mnich, Stephen J; Nemirovskiy, Olga V; Carter, Jeff; Gierse, James K; Wang, Jane L; Vazquez, Michael L; Moore, William M; Masferrer, Jaime L

2010-05-15

176

Discovery of atrop fixed alkoxy-aminobenzhydrol derivatives: novel, highly potent and orally efficacious squalene synthase inhibitors.  

PubMed

We have recently reported the discovery of the new benzhydrol template, which has a highly potent inhibitory activity for squalene synthase, as typified by compound 1 (SSI IC(50)=0.85 nM). However, it was composed of a pair of easy rotatable atropisomers. In the effort to fix the isomerization, a highly potent alkoxy-aminobenzhydrol scaffold was developed. Some of these acquired compounds demonstrating strong cholesterol synthesis inhibitory activities in a rat hepatic cell. Moreover, two of the series compounds exhibited specific plasma lipid-lowering effects in in vivo animal models. PMID:21802309

Ichikawa, Masanori; Yokomizo, Aki; Itoh, Masao; Haginoya, Noriyasu; Sugita, Kazuyuki; Usui, Hiroyuki; Terayama, Koji; Kanda, Akira

2011-09-01

177

Preparation of chitin nanogels containing nickel nanoparticles.  

PubMed

In this work, we developed 120-150 nm sized nickel nanoparticles loaded chitin nanogels (Ni-Chitin NGs) by regeneration chemistry approach to investigate and determine its cytocompatibility and antibacterial activity against Staphylococcus aureus. The nickel nanoparticles were prepared by hydrothermal method. The prepared Ni-Chitin NGs were well characterized by SEM, FTIR, TG/DTA/DTG and XRD and the in vitro cytocompatibility was tested on A549 and L929 cells which showed that they are completely non-toxic. Ni-Chitin NGs showed better toxicity to the bacterial strains when compared to previous study with other nanoparticles using serial dilution method. The rhodamine labeled-Ni-Chitin NGs showed cellular localization on both L929 and A549 cells without perturbing their cellular constituents. These studies showed that the Ni-Chitin NGs could be used for various applications in biomedical filed. PMID:23911472

Kumar, N Ashwin; Rejinold, N Sanoj; Anjali, P; Balakrishnan, Avinash; Biswas, Raja; Jayakumar, R

2013-09-12

178

Utilisation of chitinous materials in pigment adsorption.  

PubMed

The effect of adding the cells of four lactobacilli to a squid pen powder (SPP)-containing medium on prodigiosin (PG) production by Serratia marcescens TKU011 is examined. The best increase in PG productivity was shown by strain TKU012. Among the samples of strain TKU012 and the chitinous materials of cicada casting powder (CCP), shrimp shell powder (SSP), squid pen powder (SPP), ?-chitin, and ?-chitin, TKU012 cells displayed the best adsorption rate (84%) for PG, followed by CCP, SSP, SPP, ?-chitin, and ?-chitin. As for the water-soluble food colourants, Allura Red AC (R40) and Tartrazne (Y4), SPP and SSP had better adsorptive powers than pure chitin preparations, strain TKU012, and CCP. Treatment with organic solvents, hot alkali, or proteases (papain, bromelain) diminished the adsorption rates of the biosorbents. PMID:22953835

Wang, San-Lang; Chen, Yan-Cheng; Yen, Yue-Horng; Liang, Tzu-Wen

2012-12-01

179

Design, synthesis and antibacterial activities of vanillic acylhydrazone derivatives as potential ?-ketoacyl-acyl carrier protein synthase III (FabH) inhibitors.  

PubMed

Fatty acid biosynthesis is essential for bacterial survival. FabH, ?-ketoacyl-acyl carrier protein (ACP) synthase III, is a particularly attractive target, since it is central to the initiation of fatty acid biosynthesis and is highly conserved among Gram-positive and Gram-negative bacteria. A series of acylhydrazone derivatives were synthesized and developed as potent inhibitors of FabH. This inhibitor class demonstrates strong broad-spectrum antibacterial activity. Compounds with potent antibacterial activities were tested for their Escherichia coli FabH inhibitory activity. Especially, compound E9 showed the most potent antibacterial activity with MIC values of 0.39-1.56 ?g/mL against the tested bacterial strains and exhibited the most potent E. coli FabH inhibitory activity with IC(50) of 2.5 ?M. Docking simulation was performed to position compound E9 into the E. coli FabH active site to determine the probable binding conformation. PMID:23124163

Wang, Xiao-Liang; Zhang, Yan-Bin; Tang, Jian-Feng; Yang, Yu-Shun; Chen, Ruo-Qi; Zhang, Fei; Zhu, Hai-Liang

2012-11-01

180

Structure-Based Design of Novel Pyrimido[4,5-c]pyridazine Derivatives as Dihydropteroate Synthase Inhibitors with Increased Affinity  

SciTech Connect

Dihydropteroate synthase (DHPS) is the validated drug target for sulfonamide antimicrobial therapy. However, due to widespread drug resistance and poor tolerance, the use of sulfonamide antibiotics is now limited. The pterin binding pocket in DHPS is highly conserved and is distinct from the sulfonamide binding site. It therefore represents an attractive alternative target for the design of novel antibacterial agents. We previously carried out the structural characterization of a known pyridazine inhibitor in the Bacillus anthracis DHPS pterin site and identified a number of unfavorable interactions that appear to compromise binding. With this structural information, a series of 4,5-dioxo-1,4,5,6-tetrahydropyrimido[4,5-c]pyridazines were designed to improve binding affinity. Most importantly, the N-methyl ring substitution was removed to improve binding within the pterin pocket, and the length of the side chain carboxylic acid was optimized to fully engage the pyrophosphate binding site. These inhibitors were synthesized and evaluated by an enzyme activity assay, X-ray crystallography, isothermal calorimetry, and surface plasmon resonance to obtain a comprehensive understanding of the binding interactions from structural, kinetic, and thermodynamic perspectives. This study clearly demonstrates that compounds lacking the N-methyl substitution exhibit increased inhibition of DHPS, but the beneficial effects of optimizing the side chain length are less apparent.

Zhao, Ying; Hammoudeh, Dalia; Yun, Mi-Kyung; Qi, Jianjun; White, Stephen W.; Lee, Richard E. (Tennessee-HSC); (SJCH)

2012-05-29

181

The marine natural-derived inhibitors of glycogen synthase kinase-3? phenylmethylene hydantoins: In vitro and in vivo activities and pharmacophore modeling  

PubMed Central

The Red Sea sponge Hemimycale arabica afforded the known (Z)-5-(4-hydroxybenzylidene)-hydantoin (1). This natural phenylmethylene hydantoin (PMH) 1 and the synthetic (Z)-5-(4-(ethylthio)benzylidene)-hydantoin (2) showed potent in vitro and in vivo anti-growth and anti-invasive properties against PC-3M prostate cancer cells in MTT, spheroid disaggregation, and in mice models. To explore a possible molecular target of PMHs, the most potent synthetic analogue 2 has been virtually screened against various protein kinases. Molecular modeling study has shown that 2 can be successfully docked within the binding pocket of glycogen synthase kinase-3beta (GSK-3?) similar to the well-known GSK-3? inhibitor I-5. Several PMHs showed potent in vitro GSK-3? inhibitory activity with an IC50 range of 4–20 µM. The most potent analogue 3 showed a significant increase in liver glycogen level at the 5, 15, and 25 mg/kg dose levels, in vivo. Pharmacophore model was built and validated using in-house database of active and inactive GSK-3? inhibitors. The GSK-3? inhibitory activity of PMHs entitles them to be potential leads for the treatment of cancer, Alzheimer’s disease, bipolar disorders, stroke, different tau pathologies, and type-2 diabetes. PMID:19616957

Khanfar, Mohammad A.; Asal, Bilal Abu; Mudit, Mudit; Kaddoumi, Amal; El Sayed, Khalid A.

2009-01-01

182

Chitin, Chitinase Responses, and Invasive Fungal Infections  

PubMed Central

The human immune system is capable of recognizing and degrading chitin, an important cell wall component of pathogenic fungi. In the context of host-immune responses to fungal infections, herein we review the particular contributions and interplay of fungus and chitin recognition, and chitin-degrading enzymes, known as chitinases. The mechanisms of host chitinase responses may have implications for diagnostic assays as well as novel therapeutic approaches for patients that are at risk of contracting fatal fungal infections. PMID:22187561

Vega, Karina; Kalkum, Markus

2012-01-01

183

Differentiations of chitin content and surface morphologies of chitins extracted from male and female grasshopper species.  

PubMed

In this study, we used Fourier transform infrared spectroscopy (FT-IR), elemental analysis (EA), thermogravimetric analysis (TGA), X-ray diffractometry (XRD), and scanning electron microscopy (SEM) to investigate chitin structure isolated from both sexes of four grasshopper species. FT-IR, EA, XRD, and TGA showed that the chitin was in the alpha form. With respect to gender, two main differences were observed. First, we observed that the quantity of chitin was greater in males than in females and the dry weight of chitin between species ranged from 4.71% to 11.84%. Second, using SEM, we observed that the male chitin surface structure contained 25 - 90nm wide nanofibers and 90 - 250 nm nanopores, while no pores or nanofibers were observed in the chitin surface structure of the majority of females (nanofibers were observed only in M. desertus females). In contrast, the elemental analysis, thermal properties, and crystalline index values for chitin were similar in males and females. Also, we carried out enzymatic digestion of the isolated chitins using commercial chitinase from Streptomyces griseus. We observed that there were no big differences in digestion rate of the chitins from both sexes and commercial chitin. The digestion rates were for grasshoppers' chitins; 88.45-95.48% and for commercial chitin; 94.95%. PMID:25635814

Kaya, Murat; Lelešius, Evaldas; Nagrockait?, Radvil?; Sargin, Idris; Arslan, Gulsin; Mol, Abbas; Baran, Talat; Can, Esra; Bitim, Betul

2015-01-01

184

Differentiations of Chitin Content and Surface Morphologies of Chitins Extracted from Male and Female Grasshopper Species  

PubMed Central

In this study, we used Fourier transform infrared spectroscopy (FT-IR), elemental analysis (EA), thermogravimetric analysis (TGA), X-ray diffractometry (XRD), and scanning electron microscopy (SEM) to investigate chitin structure isolated from both sexes of four grasshopper species. FT-IR, EA, XRD, and TGA showed that the chitin was in the alpha form. With respect to gender, two main differences were observed. First, we observed that the quantity of chitin was greater in males than in females and the dry weight of chitin between species ranged from 4.71% to 11.84%. Second, using SEM, we observed that the male chitin surface structure contained 25 – 90nm wide nanofibers and 90 – 250 nm nanopores, while no pores or nanofibers were observed in the chitin surface structure of the majority of females (nanofibers were observed only in M. desertus females). In contrast, the elemental analysis, thermal properties, and crystalline index values for chitin were similar in males and females. Also, we carried out enzymatic digestion of the isolated chitins using commercial chitinase from Streptomyces griseus. We observed that there were no big differences in digestion rate of the chitins from both sexes and commercial chitin. The digestion rates were for grasshoppers’ chitins; 88.45–95.48% and for commercial chitin; 94.95%. PMID:25635814

Kaya, Murat; Lelešius, Evaldas; Nagrockait?, Radvil?; Sargin, Idris; Arslan, Gulsin; Mol, Abbas; Baran, Talat; Can, Esra; Bitim, Betul

2015-01-01

185

Structure determination of glycogen synthase kinase-3 from Leishmania major and comparative inhibitor structure?activity relationships with Trypanosoma brucei GSK-3  

SciTech Connect

Glycogen synthase kinase-3 (GSK-3) is a drug target under intense investigation in pharmaceutical companies and constitutes an attractive piggyback target for eukaryotic pathogens. Two different GSKs are found in trypanosomatids, one about 150 residues shorter than the other. GSK-3 short (GeneDB: Tb927.10.13780) has previously been validated genetically as a drug target in Trypanosoma brucei by RNAi induced growth retardation; and chemically by correlation between enzyme and in vitro growth inhibition. Here, we report investigation of the equivalent GSK-3 short enzymes of L. major (LmjF18.0270) and L. infantum (LinJ18{_}V3.0270, identical in amino acid sequences to LdonGSK-3 short) and a crystal structure of LmajGSK-3 short at 2 {angstrom} resolution. The inhibitor structure-activity relationships (SARs) of L. major and L. infantum are virtually identical, suggesting that inhibitors could be useful for both cutaneous and visceral leishmaniasis. Leishmania spp. GSK-3 short has different inhibitor SARs than TbruGSK-3 short, which can be explained mostly by two variant residues in the ATP-binding pocket. Indeed, mutating these residues in the ATP-binding site of LmajGSK-3 short to the TbruGSK-3 short equivalents results in a mutant LmajGSK-3 short enzyme with SAR more similar to that of TbruGSK-3 short. The differences between human GSK-3{beta} (HsGSK-3{beta}) and LmajGSK-3 short SAR suggest that compounds which selectively inhibit LmajGSK-3 short may be found.

Ojo, Kayode K.; Arakaki, Tracy L.; Napuli, Alberto J.; Inampudi, Krishna K.; Keyloun, Katelyn R.; Zhang, Li; Hol, Wim G.J.; Verlind, Christophe L.M.J.; Merritt, Ethan A.; Van Voorhis, Wesley C. (UWASH)

2012-04-24

186

1-Phenylsulfinyl-3-(pyridin-3-yl)naphthalen-2-ols: A new class of potent and selective aldosterone synthase inhibitors.  

PubMed

1-Phenylsulfinyl-3-(pyridin-3-yl)naphthalen-2-ols and related compounds were synthesized and evaluated for inhibition of aldosterone synthase (CYP11B2), a potential target for cardiovascular diseases associated with elevated plasma aldosterone levels like congestive heart failure and myocardial fibrosis. Introduction of substituents at the phenylsulfinyl moiety and changes of the substitution pattern at the naphthalene core were examined. Potent compounds were further examined for selectivity versus other important steroidogenic CYP enzymes, i.e. the highly homologous 11?-hydroxylase (CYP11B1), CYP17 and CYP19. The most potent compound (IC50 = 14 nM) discovered was the meta-trifluoromethoxy derivative 11, which also exhibited excellent selectivity toward CYP11B1 (SF = 415), and showed no inhibition of CYP17 and CYP19. PMID:25462268

Grombein, Cornelia M; Hu, Qingzhong; Heim, Ralf; Rau, Sabrina; Zimmer, Christina; Hartmann, Rolf W

2015-01-01

187

A Head to Head Comparison of Eneamide and Epoxyamide Inhibitors of Glucosamine-6-P Synthase from the Dapdiamide Biosynthetic Pathway  

PubMed Central

The dapdiamides are a family of antibiotics that have been presumed to be cleaved in the target cell to enzyme-inhibitory N-acyl-2,3-diaminopropionate (DAP) warheads containing two alternative electrophilic moieties. Our prior biosynthetic studies revealed that an eneamide warhead is made first and converted to an epoxyamide via a three enzyme branch pathway. Here we provide a rationale for this logic. We report that the RR-epoxyamide warhead is a more efficient covalent inactivator of glucosamine-6-phosphate synthase by an order of magnitude over the eneamide, and this difference correlates with a more than ten-fold difference in antibiotic activity for the corresponding acyl-DAP dipeptides. PMID:21520904

Hollenhorst, Marie A.; Ntai, Ioanna; Badet, Bernard; Kelleher, Neil L.; Walsh, Christopher T.

2011-01-01

188

6-(4-Pyridyl)pyrimidin-4(3H)-ones as CNS penetrant glycogen synthase kinase-3? inhibitors.  

PubMed

The discovery of a series of 6-(4-pyridyl)pyrimidin-4(3H)-ones derived from a hit compound with low molecular weight and sufficient chemical space is reported. Transformation of substituents led to subnanomolar potent inhibitors with in vivo tau phoshorylation lowering activity. PMID:24094818

Uehara, Fumiaki; Shoda, Aya; Aritomo, Keiichi; Fukunaga, Kenji; Watanabe, Kazutoshi; Ando, Ryoichi; Shinoda, Masaki; Ueno, Hiroaki; Kubodera, Hideo; Sunada, Shinji; Saito, Ken-Ichi; Kaji, Takahide; Asano, Shoichi; Eguchi, Jun-ichi; Yuki, Satoshi; Tanaka, Shinji; Yoneyama, Yukimi; Niwa, Takuro

2013-12-15

189

Inhibitor of Ovarian Cancer Cells Growth by Virtual Screening: A New Thiazole Derivative Targeting Human Thymidylate Synthase  

E-print Network

Scienze Biomediche, Metaboliche e Neuroscienze, Universita degli Studi di Modena e Reggio Emilia, Via e Reggio Emilia, Via Campi 183, 41125 Modena, Italy Molecular Discovery Limited, 215 Marsh RoadTS) was targeted through a virtual screening approach. The most optimal inhibitor identified, 2-{4-hydroxy-2

Stroud, Robert

190

16-Aza-ent-beyerane and 16-Aza-ent-trachylobane: Potent Mechanism-based Inhibitors of Recombinant ent-Kaurene Synthase from Arabidopsis thaliana1  

PubMed Central

The secondary ent-beyeran-16-yl carbocation (7) is a key branch point intermediate in mechanistic schemes to rationalize the cyclic structures of many tetra- and pentacyclic diterpenes including ent-beyerene, ent-kaurene, ent-trachylobane, and ent-atiserene, presumed precursors to > 1,000 known diterpenes. (Scheme 1) To evaluate these mechanistic hypotheses, we synthesized the heterocyclic analogues, 16-aza-ent-beyerane (12) and 16-aza-ent-trachylobane (13), by means of Hg(II)- and Pb(IV)-induced cyclizations onto the ?12 double bonds of tricyclic intermediates bearing carbamoylmethyl and aminomethyl groups at C-8. The 13,16-seco 16-nor carbamate (20a) was obtained from ent-beyeran-16-one oxime (17) by Beckmann fragmentation, hydrolysis, and Curtius rearrangement. The aza analogues inhibited recombinant ent-kaurene synthase from Arabidopsis thaliana (GST-rAtKS) with inhibition constants (IC = 1 × 10 ?7 and 1 × 10?6 50 M) similar in magnitude to the pseudo-binding constant of the bicyclic ent-copalyl diphosphate substrate (Km = 3 × 10?7 M). Large enhancements of binding affinities (IC50 = 4 × 10?9 and 2 × 10?8 M) were observed in the presence of 1 mM pyrophosphate which is consistent with a tightly bound ent-beyeranyl+/pyrophosphate? ion pair intermediate in the cyclization-rearrangement catalyzed by this diterpene synthase. The weak inhibition (IC50 = 1 × 10?5 M) exhibited by ent-beyeran-16 exo-yl diphosphate (11), and its failure to undergo bridge rearrangement to kaurene, appear to rule out the covalent diphosphate as a free intermediate. 16-Aza-ent-beyerane is proposed as an effective mimic for the ent-beyeran-16-yl carbocation with potential applications as an active site probe for the various ent- diterpene cyclases, and as a novel, selective inhibitor of gibberellin biosynthesis in plants. PMID:17892288

Roy, Arnab; Roberts, Frank G.; Wilderman, P. Ross; Zhou, Ke; Peters, Reuben J.; Coates, Robert M.

2013-01-01

191

Subfornical organ mediates pressor effect of angiotensin: Influence of nitric oxide synthase inhibitors, AT(1) and AT(2) angiotensin antagonist's receptors.  

PubMed

We investigated the influence of voltage-dependent calcium channels and nitric oxide (NO) on angiotensin II (ANG II)-pressor effect injected into subfornical organ (SFO). The influence of NO on nifedipine antipressor action has also been studied by utilizing N(W)-nitro-L-arginine methyl ester (L-NAME) (20 mug x 0.2 mul(-1)) a nitric oxide synthase inhibitor (NOSI) and 7-nitroindazole (7-NIT) (20 mug x 0.2 mul(-1)), a specific neuronal nitric oxide synthase inhibitor (nNOSI). We have also investigated the role of losartan and PD123319, selective ANG II AT(1) and AT(2) receptor nonpeptide antagonists, in the pressor effect of ANG II and in the effect of L-NAME and 7-NIT, injected into the SFO. Adult male Holtzman rats (220 to 280 g) were anesthetized with ketamine (80 mg/kg(-1) of body weight) plus xylazine (7 mg/kg(-1) of body weight), placed in a stereotaxic apparatus (David Kopf model for rats), and implanted with cannula into the SFO. Direct mean arterial blood pressure (MAP) was recorded in conscious rats in a test cage, without access to food or water. The previously implanted catheter into femoral artery was connected to a Statham (P23 Db) pressure transducer (Statham-Gould, Valley View, OH) coupled to a multichannel recorder (PowerLab Multirecord). MAP increased after ANG II injection. Pre-treatment with nifidipine (50 mug x 0.2 mul(-1) or 100 mug x 0.2 mul(-1)) followed by 25 pmol x 0.2 mul(-1) of ANG II, decreased ANG II-pressor effect. L-NAME and 7-NIT increased the elevation in MAP induced by ANG II, which was blocked by the prior injection of nifedipine. The AT(1) angiotensin antagonist losartan injected into the SFO blocked the effect of ANG II and the effects of L-NAME and 7-NIT while PD123319 did not. These results provide evidence that ANG II-pressor effect is influenced by nitrergic pathways that utilize L-type calcium channels in the SFO. PMID:20409914

Saad, Wilson Abrão; Camargo, Luiz Antonio de Arruda; Guarda, Ismael Francisco Motta Siqueira; Santos, Talmir Augusto Faria Brizola Dos

2008-01-01

192

Synthesis and SAR study of imidazoquinolines as a novel structural class of microsomal prostaglandin E? synthase-1 inhibitors.  

PubMed

The imidazoquinoline derivative 1 was found as a novel mPGES-1 inhibitor. Optimization of 1 led to the identification of the 2-chlorophenyl group at the C(2)-position and the quinolone structure at the C(4)-position. Compound 33, the most potent synthesized compound, showed excellent mPGES-1 inhibition (IC(50)=9.1nM) with high selectivity (>1000-fold) over both COX-1 and COX-2. PMID:22137787

Shiro, Tomoya; Takahashi, Hirotada; Kakiguchi, Keisuke; Inoue, Yoshifumi; Masuda, Keiki; Nagata, Hidetaka; Tobe, Masanori

2012-01-01

193

Implantable applications of chitin and chitosan  

Microsoft Academic Search

Chitin, extracted primarily from shellfish sources, is a unique biopolymer based on the N-acetyl-glucosamine monomer. More than 40 years have lapsed since this biopolymer had aroused the interest of the scientific community around the world for its potential biomedical applications. Chitin, together with its variants, especially its deacetylated counterpart chitosan, has been shown to be useful as a wound dressing

Eugene Khor; Lee Yong Lim

2003-01-01

194

A review of chitin and chitosan applications  

Microsoft Academic Search

Chitin is the most abundant natural amino polysaccharide and is estimated to be produced annually almost as much as cellulose. It has become of great interest not only as an underutilized resource, but also as a new functional material of high potential in various fields, and recent progress in chitin chemistry is quite noteworthy. The purpose of this review is

Majeti N. V Ravi Kumar

2000-01-01

195

A novel Pro197Glu substitution in acetolactate synthase (ALS) confers broad-spectrum resistance across ALS inhibitors.  

PubMed

Water chickweed (Myosoton aquaticum L.), a competitive broadleaf weed, is widespread in wheat fields in China. Tribenuron and pyroxsulam failed to control water chickweed in the same field in Qiaotian Village in 2011 and 2012, respectively. An initial tribenuron resistance confirmation test identified a resistant population (AH02). ALS gene sequencing revealed a previously unreported substitution of Glu for Pro at amino acid position 197 in resistant individuals. A purified subpopulation (WRR04) that was individually homozygous for the Pro197Glu substitution was generated and characterized in terms of its response to different classes of ALS inhibitors. A whole-plant experiment showed that the WRR04 population exhibited broad-spectrum resistance to tribenuron (SU, 318-fold), pyrithiobac sodium (PTB,?>?197-fold), pyroxsulam (TP, 81-fold), florasulam (TP,?>?36-fold) and imazethapyr (IMI, 11-fold). An in vitro ALS assay confirmed that the ALS from WRR04 showed high resistance to all the tested ALS inhibitors. These results established that the Pro197Glu substitution endows broad-spectrum resistance across ALS inhibitors in water chickweed. In addition, molecular markers were developed to rapidly identify the Pro197Glu mutation. PMID:25619909

Liu, Weitang; Yuan, Guohui; Du, Long; Guo, Wenlei; Li, Lingxu; Bi, Yaling; Wang, Jinxin

2015-01-01

196

Human stem cell osteoblastogenesis mediated by novel glycogen synthase kinase 3 inhibitors induces bone formation and a unique bone turnover biomarker profile in rats  

SciTech Connect

Wnt activation by inhibiting glycogen synthase kinase 3 (GSK-3) causes bone anabolism in rodents making GSK-3 a potential therapeutic target for osteoporotic and osteolytic metastatic bone disease. To understand the wnt pathway related to human disease translation, the ability of 3 potent inhibitors of GSK-3 (AZD2858, AR79, AZ13282107) to 1) drive osteoblast differentiation and mineralisation using human adipose-derived stem cells (hADSC) in vitro; and 2) stimulate rat bone formation in vivo was investigated. Bone anabolism/resorption was determined using clinically relevant serum biomarkers as indicators of bone turnover and bone formation assessed in femurs by histopathology and pQCT/?CT imaging. GSK-3 inhibitors caused ?-catenin stabilisation in human and rat mesenchymal stem cells, stimulated hADSC commitment towards osteoblasts and osteogenic mineralisation in vitro. AZD2858 produced time-dependent changes in serum bone turnover biomarkers and increased bone mass over 28 days exposure in rats. After 7 days, AZD2858, AR79 or AZ13282107 exposure increased the bone formation biomarker P1NP, and reduced the resorption biomarker TRAcP-5b, indicating increased bone anabolism and reduced resorption in rats. This biomarker profile was differentiated from anabolic agent PTH{sub 1–34} or the anti-resorptive Alendronate-induced changes. Increased bone formation in cortical and cancellous bone as assessed by femur histopathology supported biomarker changes. 14 day AR79 treatment increased bone mineral density and trabecular thickness, and decreased trabecular number and connectivity assessed by pQCT/?CT. GSK-3 inhibition caused hADSC osteoblastogenesis and mineralisation in vitro. Increased femur bone mass associated with changes in bone turnover biomarkers confirmed in vivo bone formation and indicated uncoupling of bone formation and resorption. - Highlights: • Wnt modulation with 3 novel GSK-3 inhibitors alters bone growth. • Human stem cell osteoblastogenesis and mineralisation produced by GSK-3 inhibition. • In rats, 3 GSK-3 inhibitors produced a unique serum bone turnover biomarker profile. • Enhanced bone formation was seen within 7 to 14 days of compound treatment in rats.

Gilmour, Peter S., E-mail: Peter.Gilmour@astrazeneca.com [New Opportunities Innovative Medicines group, AstraZeneca R and D, Alderley Park, Cheshire SK10 4TF (United Kingdom); O'Shea, Patrick J.; Fagura, Malbinder [New Opportunities Innovative Medicines group, AstraZeneca R and D, Alderley Park, Cheshire SK10 4TF (United Kingdom); Pilling, James E. [Discovery Sciences, AstraZeneca R and D, Alderley Park, Cheshire SK10 4TF (United Kingdom); Sanganee, Hitesh [New Opportunities Innovative Medicines group, AstraZeneca R and D, Alderley Park, Cheshire SK10 4TF (United Kingdom); Wada, Hiroki [R and I IMed, AstraZeneca R and D, Molndal (Sweden); Courtney, Paul F. [DMPK, AstraZeneca R and D, Alderley Park, Cheshire SK10 4TF (United Kingdom); Kavanagh, Stefan; Hall, Peter A. [Safety Assessment, AstraZeneca R and D, Alderley Park, Cheshire SK10 4TF (United Kingdom); Escott, K. Jane [New Opportunities Innovative Medicines group, AstraZeneca R and D, Alderley Park, Cheshire SK10 4TF (United Kingdom)

2013-10-15

197

Reference Genes to Study Herbicide Stress Response in Lolium sp.: Up-Regulation of P450 Genes in Plants Resistant to Acetolactate-Synthase Inhibitors  

PubMed Central

Variation in the expression of numerous genes is at the basis of plant response to environmental stresses. Non-target-site-based resistance to herbicides (NTSR), the major threat to grass weed chemical control, is governed by a subset of the genes involved in herbicide stress response. Quantitative PCR assays allowing reliable comparison of gene expression are thus key to identify genes governing NTSR. This work aimed at identifying a set of reference genes with a stable expression to be used as an internal standard for the normalisation of quantitative PCR data in studies investigating NTSR to herbicides inhibiting acetolactate synthase (ALS) in the major grass weed Lolium sp. Gene expression stability was assessed in plants resistant or sensitive to two ALS inhibitors, subjected or not to herbicide stress. Using three complementary approaches implemented in the programs BestKeeper, NormFinder and geNorm, cap-binding protein, glyceraldehyde-3-phosphate-dehydrogenase and ubiquitin were identified as the most suitable reference genes. This reference gene set can probably be used to study herbicide response in other weed species. It was used to compare the expression of the genes encoding two herbicide target enzymes (ALS and acetyl-coenzyme A carboxylase) and five cytochromes P450 (CYP) with potential herbicide-degrading activity between plants resistant or sensitive to ALS inhibitors. Overall, herbicide application enhanced CYP gene expression. Constitutive up-regulation of all CYP genes observed in resistant plants compared to sensitive plants suggested enhanced secondary metabolism in the resistant plants. Comprehensive transcriptome studies associated to gene expression analyses using the reference gene set validated here are required to unravel NTSR genetic determinants. PMID:23696834

Duhoux, Arnaud; Délye, Christophe

2013-01-01

198

Identification and in vitro evaluation of new leads as selective and competitive glycogen synthase kinase-3? inhibitors through ligand and structure based drug design.  

PubMed

Glycogen synthase kinase-3? elicits multi-functional effects on intracellular signaling pathways, thereby making the kinase a therapeutic target in multiple pathologies. Hence, it is important to selectively inhibit GSK-3? over structurally and biologically similar targets, such as CDK5. The current study was designed to identify and evaluate novel ATP-competitive GSK-3? inhibitors. The study was designed to identify new leads by ligand based drug design, structure based drug design and in vitro evaluation. The best validated pharmacophore model (AADRRR) identified using LBDD was derived from a dataset of 135 molecules. There were 357 primary hits within the SPECS database using this pharmacophore model. A SBDD approach to the GSK-3? and CDK5 proteins was applied to all primary hits, and 5 selective inhibitors were identified for GSK-3?. GSK-3? and CDK5 in vitro kinase inhibition assays were performed with these molecules to confirm their selectivity for GSK-3?. The molecules showed IC50 values ranging from 0.825?M to 1.116?M and were 23- to 57-fold selective for GSK-3?. Of all the molecules, molecule 3 had the lowest IC50 value of 0.825?M. Our research identified molecules possessing benzothiophene, isoquinoline, thiazolidinedione imidazo-isoquinoline and quinazolinone scaffolds. Potency of these molecules may be due to H-bond interaction with backbone residues of Val135, Asp133 and side chain interaction with Tyr134. Selectivity over CDK5 may be due to side chain interactions with Asp200, backbone of Val61, ionic interaction with Lys60 and ?-cationic interaction with Arg141. These selective molecules were also exhibited small atom hydrophobicity and H-bond interaction with water molecule. PMID:25064440

Darshit, B S; Balaji, B; Rani, P; Ramanathan, M

2014-09-01

199

Mechanisms of acquired resistance to the quinazoline thymidylate synthase inhibitor ZD1694 (Tomudex) in one mouse and three human cell lines.  

PubMed Central

Four cell lines, the mouse L1210 leukaemia, the human W1L2 lymphoblastoid and two human ovarian (CH1 and 41M) cell lines, were made resistant to ZD1694 (Tomudex) by continual exposure to incremental doses of the drug. A 500-fold increase in thymidylate synthase (TS) activity is the primary mechanism of resistance to ZD1694 in the W1L2:RD1694 cell line, which is consequently highly cross-resistant to other folate-based TS inhibitors, including BW1843U89, LY231514 and AG337, but sensitive to antifolates with other enzyme targets. The CH1:RD1694 cell line is 14-fold resistant to ZD1694, largely accounted for by the 4.2-fold increase in TS activity. Cross-resistance was observed to other TS inhibitors, including 5-fluorodeoxyuridine (FdUrd). 41M:RD1694 cells, when exposed to 0.1 microM [3H]ZD1694, accumulated approximately 20-fold less 3H-labelled material over 24 h than the parental line. Data are consistent with this being the result of impaired transport of the drug via the reduced folate/methotrexate carrier. Resistance was therefore observed to methotrexate but not to CB3717, a compound known to use this transport mechanism poorly. The mouse L1210:RD1694 cell line does not accumulate ZD1694 or Methotrexate (MTX) polyglutamates. Folylpolyglutamate synthetase substrate activity (using ZD1694 as the substrate) was decreased to approximately 13% of that observed in the parental line. Cross-resistance was found to those compounds known to be active through polyglutamation. PMID:7537518

Jackman, A. L.; Kelland, L. R.; Kimbell, R.; Brown, M.; Gibson, W.; Aherne, G. W.; Hardcastle, A.; Boyle, F. T.

1995-01-01

200

Protection from inorganic mercury effects on the in vivo dopamine release by ionotropic glutamate receptor antagonists and nitric oxide synthase inhibitors.  

PubMed

The possible role of ionotropics glutamate receptors on the HgCl(2)-induced dopamine (DA) release from rat striatum was investigated by using in vivo brain microdialysis technique after administration of selective NMDA and AMPA/Kainate receptors antagonists dizocilpine (MK-801), D (-)-2-amino-5-phoshonopentanoic acid (AP5), and 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX). Moreover, we have also studied the effects of nitric oxide synthase (NOS) inhibitors L-nitro-arginine methyl ester (L-NAME) and 7-nitro-indazol (7-NI) on HgCl(2)-induced DA release. Intraestriatal infusion of 1mM HgCl(2) increased striatal DA to 1717.2+/-375.4% respect to basal levels. Infusion of 1mM HgCl(2) in 400 microM MK-801 pre-treated animals produced an increase on striatal DA levels 61% smaller than that induced in non-pre-treated animals. In the case of AP5, this treatment reduced 92% the increase produced by HgCl(2) as compared to non-pre-treated rats. Nevertheless, the administration of CNQX did not produce any effect on HgCl(2)-induced dopamine release. Intrastriatal infusion of 1mM HgCl(2) in 100 microM L-NAME pre-treated animals produced an increase on extracellular DA levels 82% smaller than produced by HgCl(2) alone. In addition, the pre-treatment with 7-NI reduced 90% the increase produced by infusion of HgCl(2) alone in rats. Thus, HgCl(2)-induced DA release could be produced at last in part, by overstimulation of NMDA receptors with NO production, since administration of NMDA receptor antagonists and NOS inhibitors protected against HgCl(2) effects on DA release. PMID:17624650

Vidal, Lucía; Durán, Rafael; Faro, Lilian F; Campos, Francisco; Cervantes, Rosa C; Alfonso, Miguel

2007-09-01

201

Trehalose-6-phosphate synthase from the cat flea Ctenocephalides felis and Drosophila melanogaster: gene identification, cloning, heterologous functional expression and identification of inhibitors by high throughput screening.  

PubMed

Trehalose phosphate synthase (EC 2.4.1.15; TPS) is the crucial enzyme for the biosynthesis of trehalose, the main haemolymph sugar of insects, and therefore a potential insecticidal molecular target. In this study, we report the functional heterologous expression of Drosophila melanogaster TPS, the gene identification, full length cDNA cloning and functional expression of cat flea (Ctenocephalides felis) TPS, and the Michaelis-Menten constants for their specific substrates glucose-6-phosphate and uridinediphosphate-glucose. A novel high throughput screening-compatible TPS assay and its use for the identification of the first potent insect TPS inhibitors from a large synthetic compound collection (>115 000 compounds) is described. One compound class that emerged in this screening, the 4-substituted 2,6-diamino-3,5-dicyano-4H-thiopyrans, was further investigated by analysing preliminary structure-activity relationships. Here, compounds were identified that show low µM to high nM half maximal inhibitory concentrations on insect TPS and that may serve as lead compounds for the development of insecticides with a novel mode of action. PMID:22762304

Kern, C; Wolf, C; Bender, F; Berger, M; Noack, S; Schmalz, S; Ilg, T

2012-08-01

202

A quantitative structure-activity relationship (QSAR) study on a few series of potent, highly selective inhibitors of nitric oxide synthase.  

PubMed

QSAR study was performed on a series of 1,2-dihydro-4-quinazolinamines, 4,5-dialkylsubstituted-2-imino-1,3-thiazolidine derivatives and 4,5-disubstituted-1,3-oxazolidin-2-imine derivatives studied by Tinker et al. [J Med Chem (2003), 46, 913-916], Ueda et al. [Bioorg Med Chem (2004) 12, 4101-4116] and Ueda et al. [Bioorg Med Chem Lett (2004) 14, 313-316], respectively, as potent, highly selective inhibitors of inducible nitric oxide synthase (iNOS). The iNOS inhibition activity of the whole series of compounds was analyzed in relation to the physicochemical and molecular properties of the compounds. The QSAR analysis revealed that the inhibition potency of the compounds was controlled by a topological parameter 1chi(v) (Kier's first order valence molecular connectivity index), density (D), surface tension (St) and length (steric parameter) of a substituent. This suggested that the drug-receptor interaction predominantly involved the dispersion interaction, but the bulky molecule would face steric problem because of which the molecule may not completely fit in active sites of the receptor and thus may not have the optimum interaction. PMID:24791414

Bharti, Vishwa Deepak; Gupta, Satya P; Kumar, Harish

2014-02-01

203

In vitro activity of a new oral glucan synthase inhibitor (MK-3118) tested against Aspergillus spp. by CLSI and EUCAST broth microdilution methods.  

PubMed

MK-3118, a glucan synthase inhibitor derived from enfumafungin, and comparator agents were tested against 71 Aspergillus spp., including itraconazole-resistant strains (MIC, ? 4 ?g/ml), using CLSI and EUCAST reference broth microdilution methods. The CLSI 90% minimum effective concentration (MEC(90))/MIC(90) values (?g/ml) for MK-3118, amphotericin B, and caspofungin, respectively, were as follows: 0.12, 2, and 0.03 for Aspergillus flavus species complex (SC); 0.25, 2, and 0.06 for Aspergillus fumigatus SC; 0.12, 2, and 0.06 for Aspergillus terreus SC; and 0.06, 1, and 0.03 for Aspergillus niger SC. Essential agreement between the values found by CLSI and EUCAST (± 2 log(2) dilution steps) was 94.3%. MK-3118 was determined to be a potent agent regardless of the in vitro method applied, with excellent activity against contemporary wild-type and itraconazole-resistant strains of Aspergillus spp. PMID:23229479

Pfaller, Michael A; Messer, Shawn A; Motyl, Mary R; Jones, Ronald N; Castanheira, Mariana

2013-02-01

204

Heteroatom insertion into 3,4-dihydro-1H-quinolin-2-ones leads to potent and selective inhibitors of human and rat aldosterone synthase.  

PubMed

Aldosterone synthase (CYP11B2) catalyzes the conversion of 11-deoxycorticosterone to aldosterone via corticosterone and 18-hydroxycorticosterone. CYP11B2 is regarded as a new target for several cardiovascular diseases which are associated with chronically elevated aldosterone levels such as hypertension, congestive heart failure and myocardial fibrosis. In this paper, we optimized heterocycle substituted 3,4-dihydropyridin-2(1H)-ones as CYP11B inhibitors by systematic introduction of heteroatoms and by bioisosteric exchange of the lactame moiety by a sultame moiety. The most promising compounds regarding inhibition of human CYP11B2 and selectivity versus human enzymes CYP11B1, CYP17, and CYP19 were tested for inhibition of rat CYP11B2. Thus, we discovered compounds 4 and 9 which show potent inhibition of hCYP11B2 (IC50 < 1 nM) and the corresponding rat enzyme (4: 64%, 9: 51% inhibition, at 2 ?M). PMID:25528333

Grombein, Cornelia M; Hu, Qingzhong; Rau, Sabrina; Zimmer, Christina; Hartmann, Rolf W

2015-01-27

205

Characterization of the novel nitric oxide synthase inhibitor 7-nitro indazole and related indazoles: antinociceptive and cardiovascular effects.  

PubMed Central

1. 7-Nitro indazole (7-NI, 10-50 mg kg-1), 6-nitro indazole and indazole (25-100 mg kg-1) administered i.p. in the mouse produce dose-related antinociception in the late phase of the formalin-induced hindpaw licking and acetic acid-induced abdominal constriction assays. The ED50 values (mg kg-1) were as follows: 7-NI (27.5 and 22.5), 6-nitro indazole (62.5 and 44.0) and indazole (41.0 and 48.5) in the two assays respectively. 3-Indazolinone, 6 amino indazole and 6-sulphanilimido indazole (all 50 mg kg-1) were without effect. With the exception of 5-nitro indazole (50 mg kg-1) which produced sedation, none of the other indazole derivates examined caused overt behavioural changes. 2. The antinociceptive effect of 7-NI (25 mg kg-1, i.p.) in the late phase of the formalin-induced hindpaw licking assay was partially (46.7 +/- 16.2%, n = 18) reversed by pretreatment with L- but not D-arginine (both 50 mg kg-1, i.p.). 3. The time course of 7-NI induced antinociception in the mouse was correlated with inhibition of brain (cerebellum) nitric oxide synthase (NOS) activity. Maximum antinociceptive activity and NOS inhibition was detected 18-30 min following i.p. administration. In contrast, no antinociceptive effect or inhibition of cerebellar NOS was detected 75 min post-injection. 4. 7-NI, 6-nitro indazole, indazole, 3-indazolinone and 6-amino indazole (all 50 mg kg-1) failed to influence mean arterial pressure (MAP) over the 45 min after i.p. administration in the anaesthetized mouse.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:7693278

Moore, P. K.; Wallace, P.; Gaffen, Z.; Hart, S. L.; Babbedge, R. C.

1993-01-01

206

Attenuation of acute nitrogen mustard-induced lung injury, inflammation and fibrogenesis by a nitric oxide synthase inhibitor  

SciTech Connect

Nitrogen mustard (NM) is a toxic vesicant known to cause damage to the respiratory tract. Injury is associated with increased expression of inducible nitric oxide synthase (iNOS). In these studies we analyzed the effects of transient inhibition of iNOS using aminoguanidine (AG) on NM-induced pulmonary toxicity. Rats were treated intratracheally with 0.125 mg/kg NM or control. Bronchoalveolar lavage fluid (BAL) and lung tissue were collected 1 d–28 d later and lung injury, oxidative stress and fibrosis assessed. NM exposure resulted in progressive histopathological changes in the lung including multifocal lesions, perivascular and peribronchial edema, inflammatory cell accumulation, alveolar fibrin deposition, bronchiolization of alveolar septal walls, and fibrosis. This was correlated with trichrome staining and expression of proliferating cell nuclear antigen (PCNA). Expression of heme oxygenase (HO)-1 and manganese superoxide dismutase (Mn-SOD) was also increased in the lung following NM exposure, along with levels of protein and inflammatory cells in BAL, consistent with oxidative stress and alveolar-epithelial injury. Both classically activated proinflammatory (iNOS{sup +} and cyclooxygenase-2{sup +}) and alternatively activated profibrotic (YM-1{sup +} and galectin-3{sup +}) macrophages appeared in the lung following NM administration; this was evident within 1 d, and persisted for 28 d. AG administration (50 mg/kg, 2 ×/day, 1 d–3 d) abrogated NM-induced injury, oxidative stress and inflammation at 1 d and 3 d post exposure, with no effects at 7 d or 28 d. These findings indicate that nitric oxide generated via iNOS contributes to acute NM-induced lung toxicity, however, transient inhibition of iNOS is not sufficient to protect against pulmonary fibrosis. -- Highlights: ? Nitrogen mustard (NM) induces acute lung injury and fibrosis. ? Pulmonary toxicity is associated with increased expression of iNOS. ? Transient inhibition of iNOS attenuates acute lung injury induced by NM.

Malaviya, Rama; Venosa, Alessandro [Department of Pharmacology and Toxicology, Ernest Mario School of Pharmacy, Rutgers University, Piscataway, NJ 08854 (United States)] [Department of Pharmacology and Toxicology, Ernest Mario School of Pharmacy, Rutgers University, Piscataway, NJ 08854 (United States); Hall, LeRoy [Drug Safety Sciences, Johnson and Johnson, Raritan, NJ 08869 (United States)] [Drug Safety Sciences, Johnson and Johnson, Raritan, NJ 08869 (United States); Gow, Andrew J. [Department of Pharmacology and Toxicology, Ernest Mario School of Pharmacy, Rutgers University, Piscataway, NJ 08854 (United States)] [Department of Pharmacology and Toxicology, Ernest Mario School of Pharmacy, Rutgers University, Piscataway, NJ 08854 (United States); Sinko, Patrick J. [Department of Pharmaceutics, Ernest Mario School of Pharmacy, Rutgers University, Piscataway, NJ 08854 (United States)] [Department of Pharmaceutics, Ernest Mario School of Pharmacy, Rutgers University, Piscataway, NJ 08854 (United States); Laskin, Jeffrey D. [Department of Environmental and Occupational Medicine, UMDNJ-Robert Wood Johnson Medical School, Piscataway, NJ 08854 (United States)] [Department of Environmental and Occupational Medicine, UMDNJ-Robert Wood Johnson Medical School, Piscataway, NJ 08854 (United States); Laskin, Debra L., E-mail: laskin@eohsi.rutgers.edu [Department of Pharmacology and Toxicology, Ernest Mario School of Pharmacy, Rutgers University, Piscataway, NJ 08854 (United States)

2012-12-15

207

An inhibitor-driven study for enhancing the selectivity of indirubin derivatives towards leishmanial Glycogen Synthase Kinase-3 over leishmanial cdc2-related protein kinase 3  

PubMed Central

Background In search of new antiparasitic agents for overcoming the limitations of current leishmaniasis chemotherapy, we have previously shown that 6-bromoindirubin-3'-oxime (6BIO) and several other 6-substituted analogues of indirubin, a naturally occurring bis-indole present in mollusks and plants, displayed reverse selectivity from the respective mammalian kinases, targeting more potently the leishmanial Cyclin-Dependent Kinase-1 (CDK1) homologue [cdc2-related protein kinase 3 (LCRK3)] over leishmanial Glycogen Synthase Kinase-3 (LGSK-3). This reversal of selectivity in Leishmania parasites compared to mammalian cells makes the design of specific indirubin-based LGSK-3 inhibitors difficult. In this context, the identification of compounds bearing specific substitutions that shift indirubin inhibition towards LGSK-3, previously found to be a potential drug target, over LCRK3 is imperative for antileishmanial targeted drug discovery. Methods A new in-house indirubin library, composed of 35 compounds, initially designed to target mammalian kinases (CDKs, GSK-3), was tested against Leishmania donovani promastigotes and intracellular amastigotes using the Alamar blue assay. Indirubins with antileishmanial activity were tested against LGSK-3 and LCRK3 kinases, purified from homologous expression systems. Flow cytometry (FACS) was used to measure the DNA content for cell-cycle analysis and the mode of cell death. Comparative structural analysis of the involved kinases was then performed using the Szmap algorithm. Results We have identified 7 new indirubin analogues that are selective inhibitors of LGSK-3 over LCRK3. These new inhibitors were also found to display potent antileishmanial activity with GI50 values of <1.5 ??. Surprisingly, all the compounds that displayed enhanced selectivity towards LGSK-3, were 6BIO analogues bearing an additional 3'-bulky amino substitution, namely a piperazine or pyrrolidine ring. A comparative structural analysis of the two aforementioned leishmanial kinases was subsequently undertaken to explain and rationalize the selectivity trend determined by the in vitro binding assays. Interestingly, the latter analysis showed that selectivity could be correlated with differences in kinase solvation thermo dynamics induced by minor sequence variations of the otherwise highly similar ATP binding pockets. Conclusions In conclusion, 3'-bulky amino substituted 6-BIO derivatives, which demonstrate enhanced specificity towards LGSK-3, represent a new scaffold for targeted drug development to treat leishmaniasis. PMID:24886176

2014-01-01

208

Purification, Characterization, and Identification of Novel Inhibitors of the ?-Ketoacyl-Acyl Carrier Protein Synthase III (FabH) from Staphylococcus aureus  

PubMed Central

Staphylococcus aureus is a versatile and dangerous pathogen and one of the major causes of community-acquired and hospital-acquired infections. The rise of multidrug-resistant strains of S. aureus requires the development of new antibiotics with previously unexploited mechanisms of action, such as inhibition of the ?-ketoacyl-acyl carrier protein (ACP) synthase III (FabH). This enzyme initiates fatty acid biosynthesis in a bacterial type II fatty acid synthase, catalyzing a decarboxylative condensation between malonyl-ACP and an acyl coenzyme A (CoA) substrate and is essential for viability. We have identified only one fabH in the genome of S. aureus and have shown that it encodes a protein with 57, 40, and 34% amino acid sequence identity with the FabH proteins of Bacillus subtilis (bFabH1), Escherichia coli (ecFabH), and Mycobacterium tuberculosis (mtFabH). Additional genomic sequence analysis revealed that this S. aureus FabH (saFabH) is not mutated in certain methicillin-resistant S. aureus (MRSA) and vancomycin-resistant S. aureus (VRSA) strains. saFabH was expressed in E. coli with an N-terminal polyhistidine tag and subsequently purified by metal chelate and size exclusion chromatography. Analysis by sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed a molecular mass of 37 kDa, while gel filtration demonstrated a mass of 66.7 kDa, suggesting a noncovalent homodimeric structure for saFabH. The apparent Km for malonyl-ACP was 1.76 ± 0.40 ?M, and the enzyme was active with acetyl-CoA (kcat, 16.18 min?1; Km, 6.18 ± 0.9 ?M), butyryl-CoA (kcat, 42.90 min?1; Km, 2.32 ± 0.12 ?M), and isobutyryl-CoA (kcat, 98.0 min?1; Km, 0.32 ± 0.04 ?M). saFabH was weakly inhibited by thiolactomycin (50% inhibitory concentration [IC50], >100 ?M) yet was efficiently inhibited by two new FabH inhibitors, 5-chloro-4-phenyl-[1,2]-dithiol-3-one (IC50, 1.87 ± 0.10 ?M) and 4-phenyl-5-phenylimino-[1,2,4]dithiazolidin-3-one (IC50, 0.775 ± 0.08 ?M). PMID:11959561

He, Xin; Reynolds, Kevin A.

2002-01-01

209

Pharmacodynamic Target Evaluation of a Novel Oral Glucan Synthase Inhibitor, SCY-078 (MK-3118), Using an In Vivo Murine Invasive Candidiasis Model.  

PubMed

Echinocandins inhibit the synthesis of ?-1,3-d-glucan in Candida and are the first-line therapy in numerous clinical settings. Their use is limited by poor oral bioavailability, and they are available only as intravenous therapies. Derivatives of enfumafungin are novel orally bioavailable glucan synthase inhibitors. We performed an in vivo pharmacodynamic (PD) evaluation with a novel enfumafungin derivative, SCY-078 (formerly MK-3118), in a well-established neutropenic murine model of invasive candidiasis against C. albicans, C. glabrata, and C. parapsilosis. The SCY-078 MICs varied 8-fold. Oral doses of 3.125 to 200 mg/kg SCY-078 salt in sterile water produced peak levels of 0.04 to 2.66 ?g/ml, elimination half-lives of 5.8 to 8.5 h, areas under the concentration-time curve from 0 to 24 h (AUC0-24 h) of 0.61 to 41.10 ?g · h/ml, and AUC from 0 to infinity (AUC0-?) values of 0.68 to 40.31 ?g · h/ml. The pharmacokinetics (PK) were approximately linear over the dose range studied. Maximum response (Emax) and PK/PD target identification studies were performed with 4 C. albicans, 4 C. glabrata, and 3 C. parapsilosis isolates. The PD index AUC/MIC was explored by using total (tAUC) and free (fAUC) drug concentrations. The maximum responses were 4.0, 4.0, and 4.3 log10 CFU/kidney reductions for C. albicans, C. glabrata, and C. parapsilosis, respectively. The AUC/MIC was a robust predictor of efficacy (R(2), 0.53 to 0.91). The 24-h PD targets were a static dose of 63.5 mg/kg, a tAUC/MIC of 500, and an fAUC/MIC of 1.0 for C. albicans; a static dose of 58.4 mg/kg, a tAUC/MIC of 315, and an fAUC/MIC of 0.63 for C. glabrata; and a static dose of 84.4 mg/kg, a tAUC/MIC of 198, and an fAUC/MIC of 0.40 for C. parapsilosis. The mean fAUC/MIC values associated with a 1-log kill endpoint against these species were 1.42, 1.26, and 0.91 for C. albicans, C. glabrata, and C. parapsilosis, respectively. The static and 1-log kill endpoints were measured relative to the burden at the start of therapy. The static and 1-log kill doses, as well as the total and free drug AUC/MIC PD targets, were not statistically different between species but were numerically lower than those observed for echinocandins. SCY-078 is a promising novel oral glucan synthase inhibitor against Candida species, and further investigation is warranted. PMID:25512406

Lepak, Alexander J; Marchillo, Karen; Andes, David R

2015-02-01

210

Reduction of carrageenin oedema and the associated c-Fos expression in the rat lumbar spinal cord by nitric oxide synthase inhibitor.  

PubMed Central

1. Three hours after intraplantar carrageenin (6 mg/150 microliters of saline) Fos-like immunoreactivity (Fos-LI) was mainly observed in L4 and L5 segments of the dorsal horn. Both superficial (I-II) and deep laminae (V-VI) neurones were labelled. 2. We have studied the effect of systemic administration of a nitric oxide synthase inhibitor, NG-nitro-L-arginine methyl ester (L-NAME) on carrageenin evoked c-Fos expression and thus the contribution of nitric oxide to this expression. 3. Pre-administration of L-NAME (10, 25, 50, 100 mg kg-1, i.v.) dose-dependently reduced the number of superificial and deep laminae Fos-LI neurones, 100 mg kg-1 produced a 63 +/- 2% and 72 +/- 4% reduction of Fos-LI neurones respectively, P < 0.0001 for both superficial and deep neurones. 4. Pre-administered L-NAME dose-relatedly reduced the carrageenin-evoked paw and ankle oedema, with 100 mg kg-1 of L-NAME resulting in a 74 +/- 2% and 103 +/- 2% reduction respectively. 5. Post-administration of L-NAME (10 mg kg-1, i.v.) reduced the number of superficial and deep laminae Fos-LI neurones (65 +/- 7% and 53 +/- 8% reduction respectively, P < 0.01 for both superficial and deep neurones). 6. Post-administered L-NAME reduced both the paw and ankle oedema (52 +/- 8% and 62 +/- 10% reduction respectively, P < 0.0001 for both paw and ankle).(ABSTRACT TRUNCATED AT 250 WORDS) PMID:7536097

Honoré, P; Chapman, V; Buritova, J; Besson, J M

1995-01-01

211

Effects of soluble epoxide hydrolase inhibitor on the expression of fatty acid synthase in peripheral blood mononuclear cell in patients with acute coronary syndrome  

PubMed Central

Background Researches have shown that soluble epoxide hydrolase inhibitors (sEHi) can protect against the development of atherosclerosis. Simultaneously, emerging evidences have implicated the association between fatty acid synthase (FAS) and acute coronary syndrome (ACS). We tested the hypothesis that sEHi could reduce the occurrence of ACS by regulating FAS. Methods Hospitalized ACS patients were selected as the ACS group (n = 65) while healthy normal subjects as the control group (n = 65). The blood levels of lipoproteins, fasting glucose, myocardial enzyme and high-sensitivity C-reactive protein (hs-CRP) were measured within 24 hours after admission. The peripheral blood mononuclear cells (PBMCs) were isolated and cultured. Trans-4-[4-(3-Adamantan-1-ylureido)cyclohexyloxy] benzoic acid (t-AUCB), a kind of sEHi, was then added to cells in various concentrations (0, 10, 50, 100 ?mol/L). The expression of FAS, interleukin-6 (IL-6) mRNA and protein was detected by real-time PCR or Western blot, respectively. Results (1) Compared with the control group, the serum concentration of hs-CRP in the ACS group was increased (P<0.05). The expression of FAS, IL-6 mRNA and protein were significantly increased in PBMCs from the ACS group (all P<0.05). Moreover, the levels of FAS and IL-6 mRNA were positively correlated with the serum concentration of hs-CRP (r = 0.685, P<0.01; r = 0.715, P<0.01) respectively. (2) The expression of FAS, IL-6 mRNA and protein in PBMCs from the ACS group were dose-dependently inhibited by sEHi (all P<0.05). Conclusions sEH inhibition regulated FAS and inhibited inflammation in cultured PBMCs from ACS patients, a mechanism that might prevent rupture of atherosclerotic lesions and protect against development of ACS. PMID:23305094

2013-01-01

212

Nuclear Factor-?B (NF-?B) Mediates a Protective Response in Cancer Cells Treated with Inhibitors of Fatty Acid Synthase*  

PubMed Central

The efficacy of drugs used to treat cancer can be significantly attenuated by adaptive responses of neoplastic cells to drug-induced stress. To determine how cancer cells respond to inhibition of the enzyme fatty acid synthase (FAS), we focused on NF-?B-mediated pathways, which can be activated by various cellular stresses. Treating lung cancer cells with C93, a pharmacological inhibitor of FAS, results in changes indicative of a rapid initiation of NF-?B signaling, including translocation of RelA/p65 NF-?B to the nucleus, activation of a transfected NF-?B-luciferase reporter, and increased expression of NF-?B-dependent transcripts, IL-6, IL-8, and COX-2. Verifying that these responses to C93 are specifically related to inhibition of FAS, we confirmed that levels of these same transcripts increase in response to siRNA targeting FAS. Inhibiting this NF-?B response (either by transfecting a mutant I?B? or treating with bortezomib) resulted in increased cell killing by C93, indicating that the NF-?B response is protective in this setting. Because inhibiting FAS leads to accumulation of intermediate metabolites of fatty acid biosynthesis, we then questioned whether protein kinase C (PKC) is involved in this response to metabolic stress. Immunofluorescence microscopy revealed that C93 treatment results in cellular translocation of PKC? and PKC? isoforms and increased PKC?-dependent phosphorylation of the I?B? subunit of NF-?B. Furthermore, inhibiting PKC activity with RO-31–8220 or PKC? isoform-specific siRNA attenuates C93-induced I?B? phosphorylation and NF-?B activation and also potentiates C93-induced cell killing. These results suggest a link between PKC and NF-?B in protecting cancer cells from metabolic stress induced by inhibiting FAS. PMID:21768098

Lemmon, Colleen R. M.; Woo, Ju-Hyung; Tully, Ellen; Wilsbach, Kathleen; Gabrielson, Edward

2011-01-01

213

Effect of a nitric oxide synthase inhibitor NG-nitro-L-arginine methyl ester on invasion of human colorectal cancer cell line SL-174T  

PubMed Central

AIM: To investigate the effect and mechanism of action of the nitric oxide synthase (NOS) inhibitor NG-nitro-L-arginine methyl ester (L-NAME) on invasion and metastasis of human colorectal cancer cell line SL-174T. METHODS: Human colorectal cancer cell line SL-174T was cultured and treated separately with four different dosages of L-NAME for 72 h. Nitric oxide (NO) production was measured with Griess reagent. The effect of L-NAME on invasion and migration of SL-174T cells were evaluated by using Transwell chambers attached with polycarbonate filters and reconstituted basement membrane (Matrigel). RT-PCR was performed to determine the mRNA levels of matrix metalloproteinase-2 (MMP-2) and tissue inhibitor metalloproteinase-2 (TIMP-2). RESULTS: L-NAME could significantly inhibit NO production of SL-174T in a dose-dependent manner. After being treated for 72 h with 0.2, 0.4, 0.8, and 1.0 mmol/L L-NAME, respectively, the ability of the L-NAME treated SL-174T cells to invade the reconstituted basement membrane decreased significantly (t = 8.056, P<0.05; t = 14.467, P<0.01; t = 27.785, P<0.01; and t = 29.405, P<0.01, respectively) and the inhibition rates were 10.29%, 19.62%, 34.08%, and 42.23%, respectively. Moreover, L-NAME could inhibit migration of SL-174T cells, and the inhibition rates were 20.76%, 24.95%, 39.43%, and 46.85% for L-NAME at 0.2, 0.4, 0.8, and 1.0 mmol/L, respectively (t = 15.116, P<0.01). In addition, after treatment with L-NAME, expression of MMP-2 mRNA was significantly decreased (t = 71.238, P<0.01) and that of TIMP-2 mRNA was markedly increased (t = -13.020, P<0.01). CONCLUSION: L-NAME exerts anti-invasive and anti-metastatic effects on SL-174T cell line via downregulating MMP-2 mRNA expression and upregulating TIMP-2 mRNA expression. PMID:16419170

Yu, Li-Bo; Dong, Xin-Shu; Sun, Wen-Zhou; Zhao, Dong-Lu; Yang, Yue

2005-01-01

214

Chitin enhances obese inflammation ex vivo.  

PubMed

Infection has been implicated as a co-risk factor for obesity, but the mechanism remains uncertain. Elevated levels of plasma chitinase 3-like 1 (CHI3L1) are found in obese individuals. Since CHI3L1 is produced by activated immune cells including macrophages and recognizes microbial N-acetylglucosamine polymer (chitin), we asked whether the plasma CHI3L1 protein change in obese individuals might alter their innate immune response to chitin. Thirty-six subjects (15 obese and 21 non-obese), ages 18-30 years, were recruited. Peripheral blood mononuclear cells (PBMCs) were cultured with chitin microparticles (CMP; 1-10 ?m) for 24h; tumor necrosis factor ? (TNF-?), interleukin 6 (IL-6), and CHI3L1 in the culture supernatants were measured. We chose CMP, since neither large chitin beads (40-100 ?m), chitosan microparticles (1-10 ?m), nor soluble chitin induced the cytokine/CHI3L1 production by PBMCs isolated from non-obese PBMCs ex vivo. We found that the quantity of IL-6, but not TNF-? or CHI3L1, induced by CMP was significantly correlated with plasma IL-6, BMI, waist/hip circumferences, fasting plasma insulin, and insulin resistance. These findings suggest that chitin, a substrate of CHI3L1, further promotes obese inflammation in a size- and chemical composition- dependent manner. PMID:24055693

Huang, Chun-Jung; Beasley, Kathleen N; Acevedo, Edmund O; Franco, Robert L; Jones, Tamekia L; Mari, David C; Shibata, Yoshimi

2014-01-01

215

Spermine synthase  

Microsoft Academic Search

Spermine is present in many organisms including animals, plants, some fungi, some archaea, and some bacteria. It is synthesized\\u000a by spermine synthase, a highly specific aminopropyltransferase. This review describes spermine synthase structure, genetics,\\u000a and function. Structural and biochemical studies reveal that human spermine synthase is an obligate dimer. Each monomer contains\\u000a a C-terminal domain where the active site is located,

Anthony E. PeggAnthony; Anthony J. Michael

2010-01-01

216

Inhibitors of the fungal cell wall. Synthesis of 4-aryl-4- N-arylamine-1-butenes and related compounds with inhibitory activities on ?(1–3) glucan and chitin synthases  

Microsoft Academic Search

As part of our project devoted to the search for antifungal agents, which act via a selective mode of action, we synthesized a series of new 4-aryl- or 4-alkyl-N-arylamine-1-butenes and transformed some of them into 2-substituted 4-methyl-tetrahydroquinolines and quinolines by using a novel three-step synthesis. Results obtained in agar dilution assays have shown that 4-aryl homoallylamines not possessing halogen in

Juan M Urbina; Juan C. G Cortés; Alirio Palma; Silvia N López; Susana A Zacchino; Ricardo D Enriz; Juan C Ribas; Vladimir V Kouznetzov

2000-01-01

217

Non-Bisphosphonate Inhibitors of Isoprenoid Biosynthesis Identified via Computer-Aided Drug Design  

PubMed Central

The relaxed complex scheme, a virtual-screening methodology that accounts for protein receptor flexibility, was used to identify a low-micromolar, non-bisphosphonate inhibitor of farnesyl diphosphate synthase. Serendipitously, we also found that several predicted farnesyl diphosphate synthase inhibitors were low-micromolar inhibitors of undecaprenyl diphosphate synthase. These results are of interest because farnesyl diphosphate synthase inhibitors are being pursued as both anti-infective and anticancer agents, and undecaprenyl diphosphate synthase inhibitors are antibacterial drug leads. PMID:21696546

Durrant, Jacob D; Cao, Rong; Gorfe, Alemayehu A; Zhu, Wei; Li, Jikun; Sankovsky, Anna; Oldfield, Eric; McCammon, J Andrew

2011-01-01

218

Determinants of Activity of the Antifolate Thymidylate Synthase Inhibitors Tomudex (ZD1694) and GW1843U89 against Mono and Multilayered Colon Cancer Cell Lines under Folate-restricted Conditions1  

Microsoft Academic Search

The cytotoxicity and metabolic effects of two thymidylate synthase (TS) inhibitors, Tomudex (Raltitrexed, ZD1694) and GW1843U89, were stud- ied in WiDr colon cancer cells under four different growth conditions: as standard monolayers and as postconfluent multilayers grown under either high (WiDr, 8.8 mM folic acid) or low (WiDr\\/F, 1 nM leucovorin) folate conditions. Both GW1843U89 and ZD1694 were 13-15-fold more

Godefridus J. Peters; Evelien Smitskamp-Wilms; Kees Smid; Herbert M. Pinedo; Gerrit Jansen

1999-01-01

219

Synthesis, characterization and thermal properties of chitin-g-poly(epsilon-caprolactone) copolymers by using chitin gel.  

PubMed

Chitin is known to be natural polymer and it is non-toxic, biodegradable and biocompatible. The chitin-g-poly(epsilon-caprolactone) (chitin-g-PCL) copolymer was prepared by the ring-opening polymerization of epsilon-caprolactone onto chitin gel in the presence of tin(II) 2-ethylhexanoate catalyst by bulk polymerization method under homogeneous system. The prepared copolymer were characterized by FT-IR, (13)C NMR, thermogravimetric analysis (TGA), differential thermal analysis (DTA), scanning electron microscopy (SEM), solubility and X-ray diffraction (XRD). The degree of substitution of chitin-g-PCL copolymer was found to be 0.48. The TGA analysis showed that chitin-g-PCL was slightly less thermal stability than original chitin. It was due to the grafting of PCL reduced the crystalline structure of chitin. DTA analysis of chitin-g-PCL showed the two exothermic peaks between 300 and 400 degrees C. The first peak at 342 degrees C was due to chitin peak and the second peak was due to PCL. These results suggested that chitin and PCL chains were mixed well at a molecular level. The XRD pattern analysis of chitin-g-PCL showed a weak and broader peak, which demonstrated that the conjugation of PCL with chitin suppressed the crystallization of both chitin and PCL to some extent. The SEM studies showed that the chitin gel seems have a smooth surface morphology, but the chitin-g-PCL showed slightly rough morphology due to the grafting of PCL into chitin. The surface morphology studies also confirmed the grafting reaction. PMID:17950453

Jayakumar, R; Tamura, H

2008-07-01

220

Bacterial chitin degradation—mechanisms and ecophysiological strategies  

PubMed Central

Chitin is one the most abundant polymers in nature and interacts with both carbon and nitrogen cycles. Processes controlling chitin degradation are summarized in reviews published some 20 years ago, but the recent use of culture-independent molecular methods has led to a revised understanding of the ecology and biochemistry of this process and the organisms involved. This review summarizes different mechanisms and the principal steps involved in chitin degradation at a molecular level while also discussing the coupling of community composition to measured chitin hydrolysis activities and substrate uptake. Ecological consequences are then highlighted and discussed with a focus on the cross feeding associated with the different habitats that arise because of the need for extracellular hydrolysis of the chitin polymer prior to metabolic use. Principal environmental drivers of chitin degradation are identified which are likely to influence both community composition of chitin degrading bacteria and measured chitin hydrolysis activities. PMID:23785358

Beier, Sara; Bertilsson, Stefan

2013-01-01

221

From a Natural Product Lead to the Identification of Potent and Selective Benzofuran-3-yl-(indol-3-yl)maleimides as Glycogen Synthase Kinase 3? Inhibitors that Suppress Proliferation and Survival of Pancreatic Cancer Cells  

PubMed Central

Recent studies have demonstrated that Glycogen Synthase Kinase 3? (GSK-3?) is overexpressed in human colon and pancreatic carcinomas contributing to cancer cell proliferation and survival. Here, we report the design, synthesis, and biological evaluation of benzofuran-3-yl-(indol-3-yl)maleimides, potent GSK-3? inhibitors. Some of these compounds show picomolar inhibitory activity toward GSK-3? and an enhanced selectivity against Cyclin-dependent Kinase 2 (CDK-2). Selected GSK-3? inhibitors were tested in the pancreatic cancer cell lines MiaPaCa-2, BXPC-3, and HupT3. We determined that some of these compounds, namely compounds 5, 6, 11, 20 and 26, demonstrate antiproliferative activity against some or all of the pancreatic cancer cells at low micromolar to nanomolar concentrations. We found that the treatment of pancreatic cancer cells with GSK-3? inhibitors 5 and 26 resulted in suppression of GSK-3? activity and a distinct decrease of the X-linked Inhibitor of Apoptosis (XIAP) expression leading to significant apoptosis. The present data suggest a possible role for GSK-3? inhibitors in cancer therapy, in addition to their more prominent applications in CNS disorders. PMID:19338355

Gaisina, Irina N.; Gallier, Franck; Ougolkov, Andrei V.; Kim, Ki H.; Kurome, Toru; Guo, Songpo; Holzle, Denise; Luchini, Doris N.; Blond, Sylvie Y.; Billadeau, Daniel D.; Kozikowski, Alan P.

2009-01-01

222

Substrate and inhibitor-induced conformational changes in the structurally related enzymes UDP-N-acetylglucosamine enolpyruvyl transferase (MurA) and 5-enolpyruvylshikimate 3-phosphate synthase (EPSPS).  

PubMed

UDP-N-acetylglucosamine enolpyruvyl transferase (MurA) and 5-enolpyruvylshikimate 3-phosphate synthase (EPSPS) have both a unique three-dimensional topology and overall reaction mechanism in common. In the case of MurA, the substrate-free, unliganded protein exhibits an "open" conformation. Upon binding of substrates, the protein forms a much more tightly packed so-called "closed" form following an induced fit mechanism. In this closed form, the substrates are properly positioned for catalysis. On the basis of the structural and mechanistic similarities of MurA and EPSPS, a similar conformational change is likely to occur in EPSPS to generate a catalytically competent active site. However, there is currently little experimental evidence available to support the occurrence of such a conformational change in EPSPS. Using limited tryptic digestion of MurA,(1) it could be shown that formation of the "closed" conformation of MurA is accompanied by a marked increase of stability toward proteolytic degradation. Formation of the closed conformation was achieved by addition of either an excess of both substrates or the sugar nucleotide substrate in conjunction with the antibiotic fosfomycin. Analysis of the MurA tryptic fragments by MALDI-TOF mass spectrometry demonstrates that the protection of the protein in either case is caused by (1) a specific shielding of regions thereby becoming less accessible as a result of the conformational change, and (2) an unspecific overall protection of the whole protein due to an apparently reduced flexibility of the peptide backbone in the binary and ternary complexes. The establishment of methods to describe the effects of tryptic digestion on MurA under various conditions was then extended to EPSPS. Although EPSPS was found to be much more stable toward proteolysis than MurA, the presence of shikimate 3-phosphate (S3P) and the inhibitor glyphosate led to a pronounced suppression of proteolytic degradation. When unliganded EPSPS was treated with trypsin, three of the peptide fragments obtained could be identified by mass spectrometry. Two of these are located in a region corresponding to the "catalytic" loop in MurA which participates in the conformational change. This indicates a conformational change in EPSPS, similar to the one observed in MurA, leading to the protection mentioned above. Corroborating evidence was obtained using a conformational sensitive monoclonal antibody against EPSPS which showed a 20-fold reduced affinity toward the protein complexed with S3P and glyphosate as compared to the unliganded enzyme. PMID:10413459

Krekel, F; Oecking, C; Amrhein, N; Macheroux, P

1999-07-13

223

Nitric oxide donors prevent while the nitric oxide synthase inhibitor L-NAME increases arachidonic acid plus CYP2E1-dependent toxicity  

SciTech Connect

Polyunsaturated fatty acids such as arachidonic acid (AA) play an important role in alcohol-induced liver injury. AA promotes toxicity in rat hepatocytes with high levels of cytochrome P4502E1 and in HepG2 E47 cells which express CYP2E1. Nitric oxide (NO) participates in the regulation of various cell activities as well as in cytotoxic events. NO may act as a protectant against cytotoxic stress or may enhance cytotoxicity when produced at elevated concentrations. The goal of the current study was to evaluate the effect of endogenously or exogenously produced NO on AA toxicity in liver cells with high expression of CYP2E1 and assess possible mechanisms for its actions. Pyrazole-induced rat hepatocytes or HepG2 cells expressing CYP2E1 were treated with AA in the presence or absence of an inhibitor of nitric oxide synthase L-N {sup G}-Nitroarginine Methylester (L-NAME) or the NO donors S-nitroso-N-acetylpenicillamine (SNAP), and (Z)-1-[-(2-aminoethyl)-N-(2-aminoethyl)]diazen-1-ium-1,2-diolate (DETA-NONO). AA decreased cell viability from 100% to 48 {+-} 6% after treatment for 48 h. In the presence of L-NAME, viability was further lowered to 23 {+-} 5%, while, SNAP or DETA-NONO increased viability to 66 {+-} 8 or 71 {+-} 6%. The L-NAME potentiated toxicity was primarily necrotic in nature. L-NAME did not affect CYP2E1 activity or CYP2E1 content. SNAP significantly lowered CYP2E1 activity but not protein. AA treatment increased lipid peroxidation and lowered GSH levels. L-NAME potentiated while SNAP prevented these changes. Thus, L-NAME increased, while NO donors decreased AA-induced oxidative stress. Antioxidants prevented the L-NAME potentiation of AA toxicity. Damage to mitochondria by AA was shown by a decline in the mitochondrial membrane potential (MMP). L-NAME potentiated this decline in MMP in association with its increase in AA-induced oxidative stress and toxicity. NO donors decreased this decline in MMP in association with their decrease in AA-induced oxidative stress and toxicity. These results indicate that NO can be hepatoprotective against CYP2E1-dependent toxicity, preventing AA-induced oxidative stress.

Wu Defeng [Department of Pharmacology and Biological Chemistry, Box 1603, One Gustave L. Levy Place, Mount Sinai School of Medicine, New York, NY 10029 (United States); Cederbaum, Arthur [Department of Pharmacology and Biological Chemistry, Box 1603, One Gustave L. Levy Place, Mount Sinai School of Medicine, New York, NY 10029 (United States)]. E-mail: arthur.cederbaum@mssm.edu

2006-10-15

224

Preparation of acrylic grafted chitin for wound dressing application  

Microsoft Academic Search

Chitin grafted with poly(acrylic acid) (chitin–PAA) was prepared with the aim of obtaining a hydrogel characteristic for wound dressing application. The chitin–PAA films were synthesized at various acrylic acid feed contents to investigate its effect on water sorption ability. Acrylic acid (AA) was first linked to chitin, acting as the active grafting sites on the chain that was further polymerized

Siriporn Tanodekaew; Malinee Prasitsilp; Somporn Swasdison; Boonlom Thavornyutikarn; Thanawit Pothsree; Rujiporn Pateepasen

2004-01-01

225

Preparation and physical properties of chitin fatty acids esters.  

PubMed

Trifluoroacetic anhydride is an effective promoter for the preparation of chitin single- and mixed-acid esters. Complete dissolution is achieved within 30 min when powdered chitin is heated at 70 degrees C in a mixed solution of carboxylic acid(s) and trifluoroacetic anhydride. Chitin esters prepared are chitin acetate, chitin butyrate, chitin hexanoate and chitin octanoate, chitin co-acetate/butyrate, chitin co-acetate/hexanoate, chitin co-acetate/octanoate, chitin co-acetate/palmitate, each from a solution of the respective reactants. The products have degrees of O-acyl substitution in a range of DS 1-2 depending on the nature of acyl group, as analyzed by gas-liquid and high-pressure liquid chromatography. Acetic acid as a mutual component for the mixed-acid esters increases the total degree of substitution, and the acetyl substitution is close to the relative distribution in the reaction mixture for chitin co-acetate/butyrate. It is favored over hexanoate, octanoate, and palmitate. The parent molecules, as calculated by the composition of the chitin esters and their molecular weights by light-scattering spectroscopy, are 30 kDa for the smallest and 150-151 kDa for the largest. Films of these chitin derivatives when cast from solution are strong and flexible with limited extensibility. By dynamic mechanical analysis of the ester film, it was found that both the glass transition temperature (T(g)) and the tensile modulus (E' at 25 degrees C) are highest for chitin acetate (218 degrees C and 5.8 GPa), and lowest for chitin octanoate (182 degrees C and 1.5 GPa). For the other esters, these values lie between the above-cited values, where the T(g) and the E' decrease with an increase in the chain length of the acyl constituent. PMID:19091309

Yang, Byung Y; Ding, Qiong; Montgomery, Rex

2009-02-17

226

Concurrent protein synthesis is required for in vivo chitin synthesis in postmolt blue crabs  

SciTech Connect

Chitin synthesis in crustaceans involves the deposition of a protein-polysaccharide complex at the apical surface of epithelial cells which secrete the cuticle or exoskeleton. The present study involves an examination of in vivo incorporation of radiolabeled amino acids and amino sugars into the cuticle of postmolt blue crabs, Callinectes sapidus. Rates of incorporation of both 3H leucine and 3H threonine were linear with respect to time of incubation. Incorporation of 3H threonine into the endocuticle was inhibited greater than 90% in the presence of the protein synthesis inhibitor, puromycin. Linear incorporation of 14C glucosamine into the cuticle was also demonstrated; a significant improvement of radiolabeling was achieved by using 14C-N-acetylglucosamine as the labeled precursor. Incorporation of 3H-N-acetylglucosamine into the cuticle of postmolt blue crabs was inhibited 89% by puromycin, indicating that concurrent protein synthesis is required for the deposition of chitin in the blue crab. Autoradiographic analysis of control vs. puromycin-treated crabs indicates that puromycin totally blocks labeling of the new endocuticle with 3H glucosamine. These results are consistent with the notion that crustacean chitin is synthesized as a protein-polysaccharide complex. Analysis of the postmolt and intermolt blue crab cuticle indicates that the exoskeleton contains about 60% protein and 40% chitin. The predominant amino acids are arginine, glutamic acid, alanine, aspartic acid, and threonine.

Horst, M.N. (Mercer Univ., Macon, GA (USA))

1990-12-01

227

Nanofibrillar chitin aerogels as renewable base catalysts.  

PubMed

We demonstrate the fabrication of chitin nanofibril aerogels and their successful application as base catalysts for the production of useful chemicals. Squid-pen chitin nanofibrils (ChNF) with primary C2-amine groups on their crystalline surfaces were fabricated into highly porous aerogels with high specific surface areas up to 289 m(2) g(-1) using freeze-drying or a supercritical drying process. The prepared ChNF aerogel was used in the aqueous Knoevenagel-condensation reaction and acted as a highly efficient base catalyst, suggesting that the combination of the nanofibrous aerogel structure and primary C2-amines exposed on the crystalline ChNF surface was effective for continuous flow catalysis. Because the ChNF aerogel can be easily prepared from abundant and renewable chitin present in nature, this strategy is a gateway to promoting and conducting green and sustainable chemistry. PMID:25285573

Tsutsumi, Yoshiyuki; Koga, Hirotaka; Qi, Zi-Dong; Saito, Tsuguyuki; Isogai, Akira

2014-11-10

228

Spermine synthase  

PubMed Central

Spermine is present in many organisms including animals, plants, some fungi, some archaea, and some bacteria. It is synthesized by spermine synthase, a highly specific aminopropyltransferase. This review describes spermine synthase structure, genetics, and function. Structural and biochemical studies reveal that human spermine synthase is an obligate dimer. Each monomer contains a C-terminal domain where the active site is located, a central linking domain that also forms the lid of the catalytic domain, and an N-terminal domain that is structurally very similar to S-adenosylmethionine decarboxylase. Gyro mice, which have an X-chromosomal deletion including the spermine synthase (SMS) gene, lack all spermine and have a greatly reduced size, sterility, deafness, neurological abnormalities, and a tendency to sudden death. Mutations in the human SMS lead to a rise in spermidine and reduction of spermine causing Snyder-Robinson syndrome, an X-linked recessive condition characterized by mental retardation, skeletal defects, hypotonia, and movement disorders. PMID:19859664

Michael, Anthony J.

2010-01-01

229

Biodegradation of the chitin-protein complex in crustacean cuticle  

USGS Publications Warehouse

Arthropod cuticles consist predominantly of chitin cross-linked with proteins. While there is some experimental evidence that this chitin-protein complex may resist decay, the chemical changes that occur during degradation have not been investigated in detail. The stomatopod crustacean Neogonodactylus oerstedii was decayed in the laboratory under anoxic conditions. A combination of pyrolysis-gas chromatography/mass spectrometry and FTIR revealed extensive chemical changes after just 2 weeks that resulted in a cuticle composition dominated by chitin. Quantitative analysis of amino acids (by HPLC) and chitin showed that the major loss of proteins and chitin occurred between weeks 1 and 2. After 8 weeks tyrosine, tryptophan and valine are the most prominent amino acid moieties, showing their resistance to degradation. The presence of cyclic ketones in the pyrolysates indicates that mucopolysaccharides or other bound non-chitinous carbohydrates are also resistant to decay. There is no evidence of structural degradation of chitin prior to 8 weeks when FTIR revealed a reduction in chitin-specific bands. The chemical changes are paralleled by structural changes in the cuticle, which becomes an increasingly open structure consisting of loose chitinous fibres. The rapid rate of decay in the experiments suggests that where chitin and protein are preserved in fossil cuticles degradation must have been inhibited.Arthropod cuticles consist predominantly of chitin cross-linked with proteins. While there is some experimental evidence that this chitin-protein complex may resist decay, the chemical changes that occur during degradation have not been investigated in detail. The stomatopod crustacean Neogonodactylus oerstedii was decayed in the laboratory under anoxic conditions. A combination of pyrolysis-gas chromatography/mass spectrometry and FTIR revealed extensive chemical changes after just 2 weeks that resulted in a cuticle composition dominated by chitin. Quantitative analysis of amino acids (by HPLC) and chitin showed that the major loss of proteins and chitin occurred between weeks 1 and 2. After 8 weeks tyrosine, tryptophan and valine are the most prominent amino acid moieties, showing their resistance to degradation. The presence of cyclic ketones in the pyrolysates indicates that mucopolysaccharides or other bound non-chitinous carbohydrates are also resistant to decay. There is no evidence of structural degradation of chitin prior to 8 weeks when FTIR revealed a reduction in chitin-specific bands. The chemical changes are paralleled by structural changes in the cuticle, which becomes an increasingly open structure consisting of loose chitinous fibres. The rapid rate of decay in the experiments suggests, that where chitin and protein are preserved in fossil cuticles degradation must have been inhibited.

Artur, Stankiewicz B.; Mastalerz, M.; Hof, C.H.J.; Bierstedt, A.; Flannery, M.B.; Briggs, D.E.G.; Evershed, R.P.

1998-01-01

230

A Candida albicans strain with high MIC for caspofungin and no FKS1 mutations exhibits a high chitin content and mutations in two chitinase genes.  

PubMed

We studied the cell wall of a Candida albicans laboratory mutant exhibiting a high minimum inhibitory concentration (MIC; 8 ?g ml(-1)) for caspofungin without bearing FKS1 mutations. This strain showed a reduced level of ß 1,3 D glucan (0.43×) and a higher chitin content (2.3×) than a control strain even when grown without caspofungin. No significant over- or under-expression of chitin synthase or chitinase genes was observed. However, point mutations were detected in the chitinase 2 and 3 genes. These mutations, which may affect the enzymatic activity of the encoded protein products involved in the degradation of the chitin, could have led to an increased concentration of that component, allowing the strain to compensate for its low ß 1,3 D glucan content and the effect of caspofungin. PMID:21108572

Drakulovski, P; Dunyach, C; Bertout, S; Reynes, J; Mallié, M

2011-07-01

231

Chitin and chitosan: Properties and applications  

Microsoft Academic Search

Chitin is the second most important natural polymer in the world. The main sources exploited are two marine crustaceans, shrimp and crabs. Our objective is to appraise the state of the art concerning this polysaccharide: its morphology in the native solid state, methods of identification and characterization and chemical modifications, as well as the difficulties in utilizing and processing it

Marguerite Rinaudo

2006-01-01

232

Synthesis, anticandidal activity of N(3)-(4-methoxyfumaroyl)-(S)-2,3-diaminopropanoic amide derivatives - Novel inhibitors of glucosamine-6-phosphate synthase.  

PubMed

Novel FMDP amides 4-6 have been synthesized and tested against Candida strains. The anticandidal activity has been confined only to Candida albicans. Anticandidal activity of the tested amides has correlated with their inhibitory activity of glucosamine-6-phosphate synthase in cell free extract from C. albicans. PMID:25497131

Pawlak, Dorota; Stolarska, Magdalena; Wojciechowski, Marek; Andruszkiewicz, Ryszard

2015-01-27

233

Chitin: 'Forgotten' Source of Nitrogen: From Modern Chitin to Thermally Mature Kerogen: Lessons from Nitrogen Isotope Ratios  

USGS Publications Warehouse

Chitinous biomass represents a major pool of organic nitrogen in living biota and is likely to have contributed some of the fossil organic nitrogen in kerogen. We review the nitrogen isotope biogeochemistry of chitin and present preliminary results suggesting interaction between kerogen and ammonium during thermal maturation. Modern arthropod chitin may shift its nitrogen isotope ratio by a few per mil depending on the chemical method of chitin preparation, mostly because N-containing non-amino-sugar components in chemically complex chitin cannot be removed quantitatively. Acid hydrolysis of chemically complex chitin and subsequent ion-chromatographic purification of the "deacetylated chitin-monomer" D-glucosamine (in hydrochloride form) provides a chemically well-defined, pure amino-sugar substrate for reproducible, high-precision determination of ??15N values in chitin. ??15N values of chitin exhibited a variability of about one per mil within an individual's exoskeleton. The nitrogen isotope ratio differed between old and new exoskeletons by up to 4 per mil. A strong dietary influence on the ??15N value of chitin is indicated by the observation of increasing ??15N values of chitin from marine crustaceans with increasing trophic level. Partial biodegradation of exoskeletons does not significantly influence ??15N values of remaining, chemically preserved amino sugar in chitin. Diagenesis and increasing thermal maturity of sedimentary organic matter, including chitin-derived nitrogen-rich moieties, result in humic compounds much different from chitin and may significantly change bulk ??15N values. Hydrous pyrolysis of immature source rocks at 330??C in contact with 15N-enriched NH4Cl, under conditions of artificial oil generation, demonstrates the abiogenic incorporation of inorganic nitrogen into carbon-bound nitrogen in kerogen. Not all organic nitrogen in natural, thermally mature kerogen is therefore necessarily derived from original organic matter, but may partly result from reaction with ammonium-containing pore waters.

Schimmelmann, A.; Wintsch, R.P.; Lewan, M.D.; DeNiro, M.J.

1998-01-01

234

Comparison of chitin structures isolated from seven Orthoptera species.  

PubMed

Differences in the physichochemical properties of the chitin structure of the exoskeleton of seven species from four genera were investigated in this study. The same method was used to isolate the chitin structure of the seven species. The physicochemical properties of the isolated chitins were revealed by ESEM, FTIR, TGA and XRD analyses. The FTIR, TGA and XRD results from the chitin samples were similar. The surface morphologies of the chitins were investigated by ESEM and interesting results were noted. While the surface morphologies of the chitins isolated from two species within the same genus were quite different, the surface morphologies of chitins isolated from species belonging to different genera showed similarity. It was determined that the dry weight chitin contents of the grasshopper species varied between 5.3% and 8.9%. The results of molecular analysis showed that the chitins from seven Orthoptera species (between 5.2 and 6.8kDa) have low molecular weights. Considering that these invasive and harmful species are killed with insecticides and go to waste in large amounts, this study suggests that they should be collected and evaluated as an alternative chitin source. PMID:25290985

Kaya, Murat; Erdogan, Sevil; Mol, Abbas; Baran, Talat

2015-01-01

235

Characterization of genes for chitin catabolism in Haloferax mediterranei.  

PubMed

Chitin is the second most abundant natural polysaccharide after cellulose. But degradation of chitin has never been reported in haloarchaea. In this study, we revealed that Haloferax mediterranei, a metabolically versatile haloarchaeon, could utilize colloidal or powdered chitin for growth and poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) accumulation, and the gene cluster (HFX_5025-5039) for the chitin catabolism pathway was experimentally identified. First, reverse transcription polymerase chain reaction results showed that the expression of the genes encoding the four putative chitinases (ChiAHme, ChiBHme, ChiCHme, and ChiDHme, HFX_5036-5039), the LmbE-like deacetylase (DacHme, HFX_5027), and the glycosidase (GlyAHme, HFX_5029) was induced by colloidal or powdered chitin, and chiA Hme, chiB Hme, and chiC Hme were cotranscribed. Knockout of chiABC Hme or chiD Hme had a significant effect on cell growth and PHBV production when chitin was used as the sole carbon source, and the chiABCD Hme knockout mutant lost the capability to utilize chitin. Knockout of dac Hme or glyA Hme also decreased PHBV accumulation on chitin. These results suggested that ChiABCDHme, DacHme, and GlyAHme were indeed involved in chitin degradation in H. mediterranei. Additionally, the chitinase assay showed that each chitinase possessed hydrolytic activity toward colloidal or powdered chitin, and the major product of colloidal chitin hydrolysis by ChiABCDHme was diacetylchitobiose, which was likely further degraded to monosaccharides by DacHme, GlyAHme, and other related enzymes for both cell growth and PHBV biosynthesis. Taken together, this study revealed the genes and enzymes involved in chitin catabolism in haloarchaea for the first time and indicated the potential of H. mediterranei as a whole-cell biocatalyst in chitin bioconversion. PMID:23674154

Hou, Jing; Han, Jing; Cai, Lei; Zhou, Jian; Lü, Yang; Jin, Cheng; Liu, Jingfang; Xiang, Hua

2014-02-01

236

Dual Inhibitors of Thymidylate Synthase and Dihydrofolate Reductase as Antitumour Agents: Design, Synthesis and Biological Evaluation of Classical and Nonclassical Pyrrolo[2,3-d]pyrimidine Antifolates1  

PubMed Central

We designed and synthesized a classical analog N-[4-[(2-amino-6-ethyl-3,4-dihydro-4-oxo-7H-pyrrolo[2,3-d]pyrimidin-5-yl)thio]benzoyl]-L-glutamic acid (4) and thirteen nonclassical analogs 5-17 as potential dual thymidylate synthase (TS) and dihydrofolate reductase (DHFR) inhibitors and as antitumour agents. The key intermediate in their synthesis was 2-amino-6-ethyl-3,4-dihydro-4-oxo-7H-pyrrolo[2,3-d]pyrimidine, 22, to which various aryl thiols were conveniently attached at the 5-position via an oxidative addition reaction using iodine. For the classical analog 4, the ester obtained from the reaction was deprotected and coupled with diethyl-L-glutamate followed by saponification. Compound 4 was a potent dual inhibitor of human TS (IC50 = 90 nM) and human DHFR (IC50 = 420 nM). Compound 4 was not a substrate for human FPGS. Metabolite protection studies established TS as its principal target. Most of the nonclassical analogs were only inhibitors of human TS with IC50 values of 0.23-26 ?M. PMID:16451071

Gangjee, Aleem; Jain, Hiteshkumar D.; Phan, Jaclyn; Lin, Xin; Song, Xiaohong; McGuire, John J.; Kisliuk, Roy L.

2008-01-01

237

Design, Synthesis, and Biological Evaluation of Classical and Nonclassical 2-Amino-4-oxo-5-substituted-6-methylpyrrolo[3,2-d]pyrimidines as Dual Thymidylate Synthase and Dihydrofolate Reductase Inhibitors  

PubMed Central

We designed and synthesized a classical antifolate N-{4-[(2-amino-6-methyl-4-oxo-3,4-dihydro-5H-pyrrolo[3,2-d]pyrimidin-5-yl)methyl]benzoyl}-l-glutamic acid 4 and 11 nonclassical analogues 5–15 as potential dual thymidylate synthase (TS) and dihydrofolate reductase (DHFR) inhibitors. The key intermediate in the synthesis was N-(4-chloro-6-methyl-5H-pyrrolo[3,2-d]pyrimidin-2-yl)-2,2-dimethylpropanamide, 29, to which various 5-benzyl substituents were attached. For the classical analogue 4, the ester obtained from the N-benzylation reaction was deprotected and coupled with diethyl l-glutamate followed by saponification. Compound 4 was a potent dual inhibitor of human TS (IC50 = 46 nM, about 206-fold more potent than pemetrexed) and DHFR (IC50 = 120 nM, about 55-fold more potent than pemetrexed). The nonclassical analogues were marginal inhibitors of human TS, but four analogues showed potent T. gondii DHFR inhibition along with >100-fold selectivity compared to human DHFR. PMID:18072727

Gangjee, Aleem; Li, Wei; Yang, Jie; Kisliuk, Roy L.

2013-01-01

238

2-(2-Phenylmorpholin-4-yl)pyrimidin-4(3H)-ones; a new class of potent, selective and orally active glycogen synthase kinase-3? inhibitors.  

PubMed

A series of 2-(2-phenylmorpholin-4-yl)pyrimidin-4(3H)-ones was synthesized and examined for their inhibitory activity against glycogen synthase kinase-3? (GSK-3?). We found 21, 29 and 30 to possess potent in vitro GSK-3? inhibitory activity with good in vitro PK profiles. 21 demonstrated significant decrease of tau phosphorylation after oral administration in mice and excellent PK profiles. PMID:24176395

Fukunaga, Kenji; Uehara, Fumiaki; Aritomo, Keiichi; Shoda, Aya; Hiki, Shinsuke; Okuyama, Masahiro; Usui, Yoshihiro; Watanabe, Kazutoshi; Yamakoshi, Koichi; Kohara, Toshiyuki; Hanano, Tokushi; Tanaka, Hiroshi; Tsuchiya, Susumu; Sunada, Shinji; Saito, Ken-Ichi; Eguchi, Jun-ichi; Yuki, Satoshi; Asano, Shoichi; Tanaka, Shinji; Mori, Akiko; Yamagami, Keiji; Baba, Hiroshi; Horikawa, Takashi; Fujimura, Masatake

2013-12-15

239

Characterization of acetohydroxyacid synthase from Mycobacterium tuberculosis and the identification of its new inhibitor from the screening of a chemical library  

Microsoft Academic Search

Acetohydroxyacid synthase (AHAS) is a thiamin diphosphate- (ThDP-) and FAD-dependent enzyme that catalyzes the first common step in the biosynthetic pathway of the branched-amino acids such as leucine, isoleucine, and valine. The genes of AHAS from Mycobacterium tuberculosis were cloned, and overexpressed in E. coli and purified to homogeneity. The purified AHAS from M. tuberculosis is effectively inhibited by pyrazosulfuron

Kyoung-Jae Choi; Yeon Gyu Yu; Hoh Gyu Hahn; Jung-Do Choi; Moon-Young Yoon

2005-01-01

240

Isolation and characterization of chitin and chitosan from marine origin.  

PubMed

Nowadays, chitin and chitosan are produced from the shells of crabs and shrimps, and bone plate of squid in laboratory to industrial scale. Production of chitosan involved deproteinization, demineralization, and deacetylation. The characteristics of chitin and chitosan mainly depend on production processes and conditions. The characteristics of these biopolymers such as appearance of polymer, turbidity of polymer solution, degree of deacetylation, and molecular weight are of major importance on applications of these polymers. This chapter addresses the production processes and conditions to produce chitin, chitosan, and chito-oligosaccharide and methods for characterization of chitin, chitosan, and chito-oligosaccharide. PMID:25081074

Nwe, Nitar; Furuike, Tetsuya; Tamura, Hiroshi

2014-01-01

241

Benzalacetone Synthase  

PubMed Central

Benzalacetone synthase, from the medicinal plant Rheum palmatum (RpBAS), is a plant-specific chalcone synthase (CHS) superfamily of type III polyketide synthase (PKS). RpBAS catalyzes the one-step, decarboxylative condensation of 4-coumaroyl-CoA with malonyl-CoA to produce the C6–C4 benzalacetone scaffold. The X-ray crystal structures of RpBAS confirmed that the diketide-forming activity is attributable to the characteristic substitution of the conserved active-site “gatekeeper” Phe with Leu. Furthermore, the crystal structures suggested that RpBAS employs novel catalytic machinery for the thioester bond cleavage of the enzyme-bound diketide intermediate and the final decarboxylation reaction to produce benzalacetone. Finally, by exploiting the remarkable substrate tolerance and catalytic versatility of RpBAS, precursor-directed biosynthesis efficiently generated chemically and structurally divergent, unnatural novel polyketide scaffolds. These findings provided a structural basis for the functional diversity of the type III PKS enzymes. PMID:22645592

Shimokawa, Yoshihiko; Morita, Hiroyuki; Abe, Ikuro

2012-01-01

242

Benzalacetone synthase.  

PubMed

Benzalacetone synthase, from the medicinal plant Rheum palmatum (RpBAS), is a plant-specific chalcone synthase (CHS) superfamily of type III polyketide synthase (PKS). RpBAS catalyzes the one-step, decarboxylative condensation of 4-coumaroyl-CoA with malonyl-CoA to produce the C(6)-C(4) benzalacetone scaffold. The X-ray crystal structures of RpBAS confirmed that the diketide-forming activity is attributable to the characteristic substitution of the conserved active-site "gatekeeper" Phe with Leu. Furthermore, the crystal structures suggested that RpBAS employs novel catalytic machinery for the thioester bond cleavage of the enzyme-bound diketide intermediate and the final decarboxylation reaction to produce benzalacetone. Finally, by exploiting the remarkable substrate tolerance and catalytic versatility of RpBAS, precursor-directed biosynthesis efficiently generated chemically and structurally divergent, unnatural novel polyketide scaffolds. These findings provided a structural basis for the functional diversity of the type III PKS enzymes. PMID:22645592

Shimokawa, Yoshihiko; Morita, Hiroyuki; Abe, Ikuro

2012-01-01

243

Homologs of the Xenopus developmental gene DG42 are present in zebrafish and mouse and are involved in the synthesis of Nod-like chitin oligosaccharides during early embryogenesis.  

PubMed Central

The Xenopus developmental gene DG42 is expressed during early embryonic development, between the midblastula and neurulation stages. The deduced protein sequence of Xenopus DG42 shows similarity to Rhizobium Nod C, Streptococcus Has A, and fungal chitin synthases. Previously, we found that the DG42 protein made in an in vitro transcription/translation system catalyzed synthesis of an array of chitin oligosaccharides. Here we show that cell extracts from early Xenopus and zebrafish embryos also synthesize chitooligosaccharides. cDNA fragments homologous to DG42 from zebrafish and mouse were also cloned and sequenced. Expression of these homologs was similar to that described for Xenopus based on Northern and Western blot analysis. The Xenopus anti-DG42 antibody recognized a 63-kDa protein in extracts from zebrafish embryos that followed a similar developmental expression pattern to that previously described for Xenopus. The chitin oligosaccharide synthase activity found in extracts was inactivated by a specific DG42 antibody; synthesis of hyaluronic acid (HA) was not affected under the conditions tested. Other experiments demonstrate that expression of DG42 under plasmid control in mouse 3T3 cells gives rise to chitooligosaccharide synthase activity without an increase in HA synthase level. A possible relationship between our results and those of other investigators, which show stimulation of HA synthesis by DG42 in mammalian cell culture systems, is provided by structural analyses to be published elsewhere that suggest that chitin oligosaccharides are present at the reducing ends of HA chains. Since in at least one vertebrate system hyaluronic acid formation can be inhibited by a pure chitinase, it seems possible that chitin oligosaccharides serve as primers for hyaluronic acid synthesis. Images Fig. 2 Fig. 4 PMID:8643441

Semino, C E; Specht, C A; Raimondi, A; Robbins, P W

1996-01-01

244

Applying Molecular Dynamics Simulations to Identify Rarely Sampled Ligand-bound Conformational States of Undecaprenyl Pyrophosphate Synthase, an Antibacterial Target  

Microsoft Academic Search

Undecaprenyl pyrophosphate synthase is a cis-prenyltransferase enzyme, which is required for cell wall biosynthesis in bacteria. Undecaprenyl pyrophosphate synthase is an attractive target for antimicrobial therapy. We performed long molecular dynamics simulations and docking studies on undecaprenyl pyrophosphate synthase to investigate its dynamic behavior and the influence of protein flexibility on the design of undecaprenyl pyrophosphate synthase inhibitors. We also

William Sinko; Sarah Williams; Adam Van Wynsberghe; Jacob D. Durrant; Rong Cao; Eric Oldfield; J. Andrew McCammon

2012-01-01

245

Degradation and mineralization of chitin in an estuary  

SciTech Connect

A method for measuring microbial degradation and mineralization of radiolabeled native chitin is described. /sup 14/C-labeled chitin was synthesized in vivo by injecting shed blue crabs (Callinectes sapidus) with N-acetyl-D-(/sup 14/C)-glucosamine, allowing for its incorporation into the exoskeleton. Rates of chitin degradation and mineralization in estuarine water and sediments were determined as functions of temperature, inoculum source, and oxygen condition. Significant differences in rates between temperature treatments were evident. Q/sub 10/ values ranged from 1.2 to 2.5 for water and sediment, respectively. Increased incubation temperature also resulted in decreased lag times before onset of chitinoclastic bacterial growth and chitin degradation. The anaerobic pathway of chitin decomposition by chitinoclastic bacteria was examined with an emphasis on end product coupling to other bacterial types. Actively growing chitinoclastic bacterial isolates produced primarily acetate, hydrogen, and carbon dioxide in broth culture.

Boyer, J.

1987-01-01

246

Synthesis of chitin cycloalkyl ester derivatives and their physical properties.  

PubMed

A series of acylated chitin derivatives was prepared by reacting chitin in a solution of trifluoroacetic anhydride and each of the cyclopropyl, cyclobutyl, cyclopentyl, and cyclohexyl carboxylic acids. The degree of O-acyl substitution was in a range of 1.1-1.4 depending upon the nature of the cyclic acid added, as determined by FT-IR analysis. The solubility of the products in the organic solvents of DMF and THF increased with an increase in the cyclic chain length of the carboxylic acid. Thermal gravimetric analysis indicated that the products were stable up to 220 degrees C for chitin cyclopropanoate and cyclobutanoate, and 250 degrees C for chitin cyclopentanoate and cyclohexanoate. The surface morphology of the products by scanning electron microscopic analysis revealed porous and globular surface for chitin cyclobutanoate, cyclopentanoate, and cyclohexanoate, contrast to the dense and smooth organization for the cyclopropanoate. PMID:20691431

Bhatt, Lok Ranjan; Kim, Bo Mi; An, Chai Young; Lu, Chi chong; Chung, Yong Sik; Soung, Min Gyu; Park, Seok Hoon; Chai, Kyu Yun

2010-09-23

247

Chitin and chitosan nanofibers: preparation and chemical modifications.  

PubMed

Chitin nanofibers are prepared from the exoskeletons of crabs and prawns, squid pens and mushrooms by a simple mechanical treatment after a series of purification steps. The nanofibers have fine nanofiber networks with a uniform width of approximately 10 nm. The method used for chitin-nanofiber isolation is also successfully applied to the cell walls of mushrooms. Commercial chitin and chitosan powders are also easily converted into nanofibers by mechanical treatment, since these powders consist of nanofiber aggregates. Grinders and high-pressure waterjet systems are effective for disintegrating chitin into nanofibers. Acidic conditions are the key factor to facilitate mechanical fibrillation. Surface modification is an effective way to change the surface property and to endow nanofiber surface with other properties. Several modifications to the chitin NF surface are achieved, including acetylation, deacetylation, phthaloylation, naphthaloylation, maleylation, chlorination, TEMPO-mediated oxidation, and graft polymerization. Those derivatives and their properties are characterized. PMID:25393598

Ifuku, Shinsuke

2014-01-01

248

Surface characteristics of chitin-based shape memory polyurethane elastomers.  

PubMed

Shape memory polyurethanes (SMPUs) were prepared from polycaprolactone diol 4000 (PCL 4000), 1,4-butanediol (BDO), chitin, dimethylol propionic acid (DMPA), triethylamine (TEA) and 4,4'-diphenylmethane diisocyanate (MDI), and the structures of the synthesized materials were verified by infrared spectroscopy. The effects of chitin and DMPA contents in the polyurethane formulation on surface properties were investigated. DMPA provides function of making hydrophilic polyurethanes. The crystalline structure of chitin enhanced the hydrophobicity of the synthesized materials. Contact angle, water absorption, surface free energy, work of water adhesion and swelling behavior of the synthesized polyurethanes were affected by varying the DMPA and chitin contents. The interactions of the PU films with solvents on the surface were clearly related to the contents of DMPA and chitin in the final polyurethane formulation. PMID:19427176

Zia, Khalid Mahmood; Zuber, Mohammad; Barikani, Mehdi; Bhatti, Ijaz Ahmad; Khan, Mohammad Bilal

2009-09-01

249

Micro-CT Imaging of Denatured Chitin by Silver to Explore Honey Bee and Insect Pathologies  

Microsoft Academic Search

BackgroundChitin and cuticle coatings are important to the environmental and immune defense of honey bees and insect pollinators. Pesticides or environmental effects may target the biochemistry of insect chitin and cuticle coating. Denaturing of chitin involves a combination of deacetylation, intercalation, oxidation, Schweiger-peeling, and the formation of amine hydrochloride salt. The term “denatured chitin” calls attention to structural and property

Peter R. Butzloff

2011-01-01

250

The Dose-Dependent Immunoregulatory Effects of the Nitric Oxide Synthase Inhibitor NG-Nitro-L-Arginine Methyl Ester in Rats with SubAcute Peritonitis  

Microsoft Academic Search

BackgroundChronic inflammation accompanied by arginine deficiency, immune dysfunction, and excess nitric oxide (NO) production is a clinical condition found in patients with peritonitis. A previous study showed that the nonselective NOS inhibitor NG-nitro-L-arginine methyl ester (L-NAME) may facilitate the metabolism of the immune nutrient arginine without altering NO homeostasis in rats with sub-acute peritonitis. Here, we investigated the effects of

Chien-Chou Hsiao; Chien-Hsing Lee; Lon-Yen Tsao; Hui-Chen Lo

2012-01-01

251

Structural basis of chitin oligosaccharide deacetylation.  

PubMed

Cell signaling and other biological activities of chitooligosaccharides (COSs) seem to be dependent not only on the degree of polymerization, but markedly on the specific de-N-acetylation pattern. Chitin de-N-acetylases (CDAs) catalyze the hydrolysis of the acetamido group in GlcNAc residues of chitin, chitosan, and COS. A major challenge is to understand how CDAs specifically define the distribution of GlcNAc and GlcNH2 moieties in the oligomeric chain. We report the crystal structure of the Vibrio cholerae CDA in four relevant states of its catalytic cycle. The two enzyme complexes with chitobiose and chitotriose represent the first 3D structures of a CDA with its natural substrates in a productive mode for catalysis, thereby unraveling an induced-fit mechanism with a significant conformational change of a loop closing the active site. We propose that the deacetylation pattern exhibited by different CDAs is governed by critical loops that shape and differentially block accessible subsites in the binding cleft of CE4 enzymes. PMID:24810719

Andrés, Eduardo; Albesa-Jové, David; Biarnés, Xevi; Moerschbacher, Bruno M; Guerin, Marcelo E; Planas, Antoni

2014-07-01

252

Tetra- and Pentacyclic Triterpene Acids from the Ancient Anti-inflammatory Remedy Frankincense as Inhibitors of Microsomal Prostaglandin E2 Synthase-1  

PubMed Central

The microsomal prostaglandin E2 synthase (mPGES)-1 is the terminal enzyme in the biosynthesis of prostaglandin (PG)E2 from cyclooxygenase (COX)-derived PGH2. We previously found that mPGES-1 is inhibited by boswellic acids (IC50 = 3–30 ?M), which are bioactive triterpene acids present in the anti-inflammatory remedy frankincense. Here we show that besides boswellic acids, additional known triterpene acids (i.e., tircuallic, lupeolic, and roburic acids) isolated from frankincense suppress mPGES-1 with increased potencies. In particular, 3?-acetoxy-8,24-dienetirucallic acid (6) and 3?-acetoxy-7,24-dienetirucallic acid (10) inhibited mPGES-1 activity in a cell-free assay with IC50 = 0.4 ?M, each. Structure–activity relationship studies and docking simulations revealed concrete structure-related interactions with mPGES-1 and its cosubstrate glutathione. COX-1 and -2 were hardly affected by the triterpene acids (IC50 > 10 ?M). Given the crucial role of mPGES-1 in inflammation and the abundance of highly active triterpene acids in frankincence extracts, our findings provide further evidence of the anti-inflammatory potential of frankincense preparations and reveal novel, potent bioactivities of tirucallic acids, roburic acids, and lupeolic acids. PMID:24844534

2014-01-01

253

UDP-N-acetylmuramic acid (UDP-MurNAc) is a potent inhibitor of MurA (enolpyruvyl-UDP-GlcNAc synthase).  

PubMed

Purified recombinant MurA (enolpyruvyl-UDP-GlcNAc synthase) overexpressed in Escherichia coli had significant amounts of UDP-MurNAc (UDP-N-acetylmuramic acid) bound after purification. UDP-MurNAc is the product of MurB, the next enzyme in peptidoglycan biosynthesis. About 25% of MurA was complexed with UDP-MurNAc after five steps during purification that should have removed it. UDP-MurNAc isolated from MurA was identified by mass spectrometry, NMR analysis, and comparison with authentic UDP-MurNAc. Subsequent investigation showed that UDP-MurNAc bound to MurA tightly, with K(d,UDP)(-)(MurNAc) = 0.94 +/- 0.04 microM, as determined by fluorescence titrations using ANS (8-anilino-1-naphthalenesulfonate) as an exogenous fluorophore. UDP-MurNAc binding was competitive with ANS and phosphate, the second product of MurA, and it inhibited MurA. The inhibition patterns were somewhat ambiguous, likely being competitive with the substrate PEP (phosphoenolpyruvate) and either competitive or noncompetitive with respect to the substrate UDP-GlcNAc (UDP-N-acetylglucosamine). These results indicate a possible role for UDP-MurNAc in regulating the biosynthesis of nucleotide precursors of peptidoglycan through feedback inhibition. Previous studies indicated that UDP-MurNAc binding to MurA was not tight enough to be physiologically relevant; however, this was likely an artifact of the assay conditions. PMID:15751977

Mizyed, Shehadeh; Oddone, Anna; Byczynski, Bartosz; Hughes, Donald W; Berti, Paul J

2005-03-15

254

Selective peptide inhibitors of bifunctional thymidylate synthase-dihydrofolate reductase from Toxoplasma gondii provide insights into domain–domain communication and allosteric regulation  

PubMed Central

The bifunctional enzyme thymidylate synthase–dihydrofolate reductase (TS–DHFR) plays an essential role in DNA synthesis and is unique to several species of pathogenic protozoans, including the parasite Toxoplasma gondii. Infection by T. gondii causes the prevalent disease toxoplasmosis, for which TS–DHFR is a major therapeutic target. Here, we design peptides that target the dimer interface between the TS domains of bifunctional T. gondii TS–DHFR by mimicking ?-strands at the interface, revealing a previously unknown allosteric target. The current study shows that these ?-strand mimetic peptides bind to the apo-enzyme in a species-selective manner to inhibit both the TS and distal DHFR. Fluorescence spectroscopy was used to monitor conformational switching of the TS domain and demonstrate that these peptides induce a conformational change in the enzyme. Using structure-guided mutagenesis, nonconserved residues in the linker between TS and DHFR were identified that play a key role in domain–domain communication and in peptide inhibition of the DHFR domain. These studies validate allosteric inhibition of apo-TS, specifically at the TS–TS interface, as a potential target for novel, species-specific therapeutics for treating T. gondii parasitic infections and overcoming drug resistance. PMID:23813474

J Landau, Mark; Sharma, Hitesh; Anderson, Karen S

2013-01-01

255

Herbicide-resistant forms of Arabidopsis thaliana acetohydroxyacid synthase: characterization of the catalytic properties and sensitivity to inhibitors of four defined mutants.  

PubMed Central

Acetohydroxyacid synthase (AHAS) catalyses the first step in the synthesis of the branched-chain amino acids and is the target of several classes of herbicides. Four mutants (A122V, W574S, W574L and S653N) of the AHAS gene from Arabidopsis thaliana were constructed, expressed in Escherichia coli, and the enzymes were purified. Each mutant form and wild-type was characterized with respect to its catalytic properties and sensitivity to nine herbicides. Each enzyme had a pH optimum near 7.5. The specific activity varied from 13% (A122V) to 131% (W574L) of the wild-type and the Km for pyruvate of the mutants was similar to the wild-type, except for W574L where it was five-fold higher. The activation by cofactors (FAD, Mg2+ and thiamine diphosphate) was examined. A122V showed reduced affinity for all three cofactors, whereas S653N bound FAD more strongly than wild-type AHAS. Six sulphonylurea herbicides inhibited A122V to a similar degree as the wild-type but S653N showed a somewhat greater reduction in sensitivity to these compounds. In contrast, the W574 mutants were insensitive to these sulphonylureas, with increases in the Kiapp (apparent inhibition constant) of several hundred fold. All four mutants were resistant to three imidazolinone herbicides with decreases in sensitivity ranging from 100-fold to more than 1000-fold. PMID:9677339

Chang, A K; Duggleby, R G

1998-01-01

256

Tetra- and pentacyclic triterpene acids from the ancient anti-inflammatory remedy frankincense as inhibitors of microsomal prostaglandin E(2) synthase-1.  

PubMed

The microsomal prostaglandin E2 synthase (mPGES)-1 is the terminal enzyme in the biosynthesis of prostaglandin (PG)E2 from cyclooxygenase (COX)-derived PGH2. We previously found that mPGES-1 is inhibited by boswellic acids (IC50 = 3-30 ?M), which are bioactive triterpene acids present in the anti-inflammatory remedy frankincense. Here we show that besides boswellic acids, additional known triterpene acids (i.e., tircuallic, lupeolic, and roburic acids) isolated from frankincense suppress mPGES-1 with increased potencies. In particular, 3?-acetoxy-8,24-dienetirucallic acid (6) and 3?-acetoxy-7,24-dienetirucallic acid (10) inhibited mPGES-1 activity in a cell-free assay with IC50 = 0.4 ?M, each. Structure-activity relationship studies and docking simulations revealed concrete structure-related interactions with mPGES-1 and its cosubstrate glutathione. COX-1 and -2 were hardly affected by the triterpene acids (IC50 > 10 ?M). Given the crucial role of mPGES-1 in inflammation and the abundance of highly active triterpene acids in frankincence extracts, our findings provide further evidence of the anti-inflammatory potential of frankincense preparations and reveal novel, potent bioactivities of tirucallic acids, roburic acids, and lupeolic acids. PMID:24844534

Verhoff, Moritz; Seitz, Stefanie; Paul, Michael; Noha, Stefan M; Jauch, Johann; Schuster, Daniela; Werz, Oliver

2014-06-27

257

Explanation for Hydrogen Bonds of Chitin?Alcohols from Lewis Acid?Base Theories  

Microsoft Academic Search

The inverse gas chromatography (IGC) method was used to characterize the hydrogen bond energies of chitin?methanol and chitin?ethanol. Surface Lewis acid?base properties of the chitin were determined at the same time. Six solvents, trichloromethane, acetone, ether, tetrahydrofuran, methanol, and ethanol were used as polar probes for the IGC measuration. The hydrogen bond energies of chitin?methanol and chitin?ethanol were 23.3 and

Baoli Shi; Wei Zhao; Qianru Zhang

2007-01-01

258

Symbiosis with bacteria enhances the use of chitin by the springtail, Folsomia candida (Collembola)  

Microsoft Academic Search

Summary  The relationship between Folsomia candida and chitin-degrading microorganisms was studied. On chitin agar, 1010 bacteria were isolated per g faeces, and 3.8×1011 bacteria per g gut contents, 1\\/3 of them showing a clear (chitin-free) zone around the colony. The most abundant chitin-degrading\\u000a bacteria were Xanthomonas maltophilia and Curtobacterium sp. To determine the bacterial contribution in the use of chitin by

H. Borkott; H. Insam

1990-01-01

259

Concurrent production of chitin from shrimp shells and fungi.  

PubMed

Crustacean shells constitute the traditional and current commercial source of chitin. Conversely, the control of fungal fermentation processes to produce quality chitin makes fungal mycelia an attractive alternative source. Therefore, the exploitation of both of these sources to produce chitin in a concurrent process should be advantageous and is reported here. Three proteolytic Aspergillus niger (strains 0576, 0307 and 0474) were selected from a screening for protease activity from among 34 zygomycete and deuteromycete strains. When fungi and shrimp shell powder were combined in a single reactor, the release of protease by the fungi facilitated the deproteinization of shrimp-shell powder and the release of hydrolyzed proteins. The hydrolyzed proteins in turn were utilized as a nitrogen source for fungal growth, leading to a lowering of the pH of the fermentation medium, thereby further enhancing the demineralization of the shrimp-shell powder. The shrimp-shell powders and fungal mycelia were separated after fermentation and extracted for chitin with 5% LiCl/DMAc solvent. Chitin isolates from the shells were found to have a protein content of less than 5%, while chitin isolates from the three fungal mycelia strains had protein content in the range of 10-15%. The relative molecular weights as estimated by GPC for all chitin samples were in the 10(5) dalton range. All samples displayed characteristic profiles for chitin in their FTIR and solid-state NMR spectra. All chitin samples evaluated with MTT and Neutral Red assays with three commercial cell lines did not display cytotoxic effects. PMID:11376610

Teng, W L; Khor, E; Tan, T K; Lim, L Y; Tan, S C

2001-06-01

260

The molecular pharmacology and in vivo activity of 2-(4-chloro-6-(2,3-dimethylphenylamino)pyrimidin-2-ylthio)octanoic acid (YS121), a dual inhibitor of microsomal prostaglandin E2 synthase-1 and 5-lipoxygenase.  

PubMed

The microsomal prostaglandin E(2) synthase (mPGES)-1 is one of the terminal isoenzymes of prostaglandin (PG) E(2) biosynthesis. Pharmacological inhibitors of mPGES-1 are proposed as an alternative to nonsteroidal anti-inflammatory drugs. We recently presented the design and synthesis of a series of pirinixic acid derivatives that dually inhibit mPGES-1 and 5-lipoxygenase. Here, we investigated the mechanism of mPGES-1 inhibition, the selectivity profile, and the in vivo activity of alpha-(n-hexyl)-substituted pirinixic acid [YS121; 2-(4-chloro-6-(2,3-dimethylphenylamino)pyrimidin-2-ylthio)octanoic acid)] as a lead compound. In cell-free assays, YS121 inhibited human mPGES-1 in a reversible and noncompetitive manner (IC(50) = 3.4 muM), and surface plasmon resonance spectroscopy studies using purified in vitro-translated human mPGES-1 indicate direct, reversible, and specific binding to mPGES-1 (K(D) = 10-14 muM). In lipopolysaccharide-stimulated human whole blood, PGE(2) formation was concentration dependently inhibited (IC(50) = 2 muM), whereas concomitant generation of the cyclooxygenase (COX)-2-derived thromboxane B(2) and 6-keto PGF(1alpha) and the COX-1-derived 12(S)-hydroxy-5-cis-8,10-trans-heptadecatrienoic acid was not significantly reduced. In carrageenan-induced rat pleurisy, YS121 (1.5 mg/kg i.p.) blocked exudate formation and leukocyte infiltration accompanied by reduced pleural levels of PGE(2) and leukotriene B(4) but also of 6-keto PGF(1alpha). Taken together, these results indicate that YS121 is a promising inhibitor of mPGES-1 with anti-inflammatory efficiency in human whole blood as well as in vivo. PMID:19934399

Koeberle, Andreas; Rossi, Antonietta; Zettl, Heiko; Pergola, Carlo; Dehm, Friederike; Bauer, Julia; Greiner, Christine; Reckel, Sina; Hoernig, Christina; Northoff, Hinnak; Bernhard, Frank; Dötsch, Volker; Sautebin, Lidia; Schubert-Zsilavecz, Manfred; Werz, Oliver

2010-03-01

261

Design, synthesis, biological evaluation and X-ray crystal structure of novel classical 6,5,6-tricyclic benzo[4,5]thieno[2,3-d]pyrimidines as dual thymidylate synthase and dihydrofolate reductase inhibitors  

PubMed Central

Classical antifolates (4-7) with a tricyclic benzo[4,5]thieno[2,3-d]pyrimidine scaffold and a flexible and rigid benzoylglutamate were synthesized as dual thymidylate synthase (TS) and dihydrofolate reductase (DHFR) inhibitors. Oxidative aromatization of ethyl 2-amino-4-methyl-4,5,6,7-tetrahydro-1-benzothiophene-3-carboxylate (±)-9 to ethyl 2-amino-4-methyl-1-benzothiophene-3-carboxylate 10 with 10% Pd/C was a key synthetic step. Compounds with 2-CH3 substituents inhibited human (h) TS (IC50 = 0.26-0.8 ?M), but not hDHFR. Substitution of the 2-CH3 with a 2-NH2 increases hTS inhibition by more than 10-fold and also affords excellent hDHFR inhibition (IC50 = 0.09-0.1 ?M). This study shows that the tricyclic benzo[4,5]thieno[2,3-d]pyrimidine scaffold is highly conducive to single hTS or dual hTS-hDHFR inhibition depending on the 2-position substituents. The X-ray crystal structures of 6 and 7 with hDHFR reveal, for the first time, that tricyclics 6 and 7 bind with the benzo[4,5]thieno[2,3-d]pyrimidine ring in the folate binding mode with the thieno S mimicking the 4-amino of methotrexate. PMID:21550809

Zhang, Xin; Zhou, Xilin; L.Kisliuk, Roy; Piraino, Jennifer; Cody, Vivian

2011-01-01

262

Effects of the 3-hydroxy-3-methylglutaryl-CoA reductase inhibitors, atorvastatin and simvastatin, on the expression of endothelin-1 and endothelial nitric oxide synthase in vascular endothelial cells.  

PubMed Central

Endothelial dysfunction associated with atherosclerosis has been attributed to alterations in the L-arginine-nitric oxide (NO)-cGMP pathway or to an excess of endothelin-1 (ET-1). The 3-hydroxy-3-methylglutaryl coenzyme A reductase inhibitors (statins) have been shown to ameliorate endothelial function. However, the physiological basis of this observation is largely unknown. We investigated the effects of Atorvastatin and Simvastatin on the pre-proET-1 mRNA expression and ET-1 synthesis and on the endothelial NO synthase (eNOS) transcript and protein levels in bovine aortic endothelial cells. These agents inhibited pre-proET-1 mRNA expression in a concentration- and time-dependent fashion (60-70% maximum inhibition) and reduced immunoreactive ET-1 levels (25-50%). This inhibitory effect was maintained in the presence of oxidized LDL (1-50 microg/ml). No significant modification of pre-proET-1 mRNA half-life was observed. In addition, mevalonate, but not cholesterol, reversed the statin-mediated decrease of pre-proET-1 mRNA levels. eNOS mRNA expression was reduced by oxidized LDL in a dose-dependent fashion (up to 57% inhibition), whereas native LDL had no effect. Statins were able to prevent the inhibitory action exerted by oxidized LDL on eNOS mRNA and protein levels. Hence, these drugs might influence vascular tone by modulating the expression of endothelial vasoactive factors. PMID:9637705

Hernández-Perera, O; Pérez-Sala, D; Navarro-Antolín, J; Sánchez-Pascuala, R; Hernández, G; Díaz, C; Lamas, S

1998-01-01

263

Nature of isomerism of solid isothiourea salts, inhibitors of nitric oxide synthases, as studied by 1H-14N nuclear quadrupole double resonance, X-ray, and density functional theory/quantum theory of atoms in molecules.  

PubMed

Isothioureas, inhibitors of nitric oxide synthases, have been studied experimentally in solid state by nuclear quadrupole double resonance (NQDR) and X-ray methods and theoretically by the quantum theory of atoms in molecules/density functional theory. Resonance frequencies on (14)N have been detected and assigned to particular nitrogen sites in each molecule. The crystal packings of (S)-3,4-dichlorobenzyl-N-methylisothiouronium chloride with the disordered chlorine positions in benzene ring and (S)-butyloisothiouronium bromide have been resolved in X-ray diffraction studies. (14)N NQDR spectra have been found good indicators of isomer type and strength of intra- or intermolecular N-H···X (X = Cl, Br) interactions. From among all salts studied, only for (S)-2,3,4,5,6-pentabromobenzylisothiouronium chloride are both nitrogen sites equivalent, which has been explained by the slow exchange. This unique structural feature can be a key factor in the high biological activity of (S)-2,3,4,5,6-pentabromobenzylisothiouronium salts. PMID:22283980

Latosi?ska, J N; Latosi?ska, M; Seliger, J; Žagar, V; Maurin, J K; Kazimierczuk, Z

2012-02-01

264

Chitin nanocrystal-xyloglucan multilayer thin films.  

PubMed

For the first time, the adsorption of xyloglucan (XG) on chitin nanocrystals (ChiNC) surface was proved using quartz crystal microbalance with dissipation (QCM-D) and by successfully building up spin-coated assisted layer-by-layer (LbL) structures on solid substrates. Several parameters in the adsorption process, such as ChiNC concentrations (0.5-3.0 g L(-1)), number of layers, or the outmost layer material (ChiNC or XG), were investigated to better understand the fabrication process of multilayer films. The thickness of the homogeneous film increased linearly with the number of bilayers, with an average thickness per bilayer of 12.3 nm. Additionally surface morphology was studied by atomic force microscopy (AFM), which revealed an almost completely covered surface after the adsorption of ChiNC. The final structures were found to have semireflective properties capable of being tuned by adjusting the ChiNC dispersion parameters. PMID:24328307

Villares, Ana; Moreau, Céline; Capron, Isabelle; Cathala, Bernard

2014-01-13

265

Novel, potent, orally bioavailable and selective mycobacterial ATP synthase inhibitors that demonstrated activity against both replicating and non-replicating M. tuberculosis.  

PubMed

The mycobacterial F0F1-ATP synthase (ATPase) is a validated target for the development of tuberculosis (TB) therapeutics. Therefore, a series of eighteen novel compounds has been designed, synthesized and evaluated against Mycobacterium smegmatis ATPase. The observed ATPase inhibitory activities (IC50) of these compounds range between 0.36 and 5.45?M. The lead compound 9d [N-(7-chloro-2-methylquinolin-4-yl)-N-(3-((diethylamino)methyl)-4-hydroxyphenyl)-2,3-dichlorobenzenesulfonamide] with null cytotoxicity (CC50>300?g/mL) and excellent anti-mycobacterial activity and selectivity (mycobacterium ATPase IC50=0.51?M, mammalian ATPase IC50>100?M, and selectivity >200) exhibited a complete growth inhibition of replicating Mycobacterium tuberculosis H37Rv at 3.12?g/mL. In addition, it also exhibited bactericidal effect (approximately 2.4log10 reductions in CFU) in the hypoxic culture of non-replicating M. tuberculosis at 100?g/mL (32-fold of its MIC) as compared to positive control isoniazid [approximately 0.2log10 reduction in CFU at 5?g/mL (50-fold of its MIC)]. The pharmacokinetics of 9d after p.o. and IV administration in male Sprague-Dawley rats indicated its quick absorption, distribution and slow elimination. It exhibited a high volume of distribution (Vss, 0.41L/kg), moderate clearance (0.06L/h/kg), long half-life (4.2h) and low absolute bioavailability (1.72%). In the murine model system of chronic TB, 9d showed 2.12log10 reductions in CFU in both lung and spleen at 173?mol/kg dose as compared to the growth of untreated control group of Balb/C male mice infected with replicating M. tuberculosis H37Rv. The in vivo efficacy of 9d is at least double of the control drug ethambutol. These results suggest 9d as a promising candidate molecule for further preclinical evaluation against resistant TB strains. PMID:25614114

Singh, Supriya; Roy, Kuldeep K; Khan, Shaheb R; Kashyap, Vivek Kr; Sharma, Abhisheak; Jaiswal, Swati; Sharma, Sandeep K; Krishnan, Manju Yasoda; Chaturvedi, Vineeta; Lal, Jawahar; Sinha, Sudhir; Gupta, Arnab D; Srivastava, Ranjana; Saxena, Anil K

2015-02-15

266

Requirement for chitin biosynthesis in epithelial tube morphogenesis  

E-print Network

, a polymer of N-acetyl- -D-glucosamine that serves as a scaffold in the rigid extracellular matrix of insect an unexpected role in tracheal tube morphogenesis for chitin, an extracellular polymer of N-acetyl- -D-glucosamine

Krasnow, Mark A.

267

Emerging chitin and chitosan nanofibrous materials for biomedical applications  

NASA Astrophysics Data System (ADS)

Over the past several decades, we have witnessed significant progress in chitosan and chitin based nanostructured materials. The nanofibers from chitin and chitosan with appealing physical and biological features have attracted intense attention due to their excellent biological properties related to biodegradability, biocompatibility, antibacterial activity, low immunogenicity and wound healing capacity. Various methods, such as electrospinning, self-assembly, phase separation, mechanical treatment, printing, ultrasonication and chemical treatment were employed to prepare chitin and chitosan nanofibers. These nanofibrous materials have tremendous potential to be used as drug delivery systems, tissue engineering scaffolds, wound dressing materials, antimicrobial agents, and biosensors. This review article discusses the most recent progress in the preparation and application of chitin and chitosan based nanofibrous materials in biomedical fields.

Ding, Fuyuan; Deng, Hongbing; Du, Yumin; Shi, Xiaowen; Wang, Qun

2014-07-01

268

Nanostructured membranes based on native chitin nanofibers prepared by mild process.  

PubMed

Procedures for chitin nanofiber or nanocrystal extraction from Crustaceans modify the chitin structure significantly, through surface deacetylation, surface oxidation and/or molar mass degradation. Here, very mild conditions were used to disintegrate chitin fibril bundles and isolate low protein content individualized chitin nanofibers, and prepare nanostructured high-strength chitin membranes. Most of the strongly 'bound' protein was removed. The degree of acetylation, crystal structure as well as length and width of the native chitin microfibrils in the organism were successfully preserved. Atomic force microscopy and scanning transmission electron microscopy, showed chitin nanofibers with width between 3 and 4 nm. Chitin membranes were prepared by filtration of hydrocolloidal nanofiber suspensions. Mechanical and optical properties were measured. The highest data so far reported for nanostructured chitin membranes was obtained for ultimate tensile strength, strain to failure and work to fracture. Strong correlation was observed between low residual protein content and high tensile properties and the reasons for this are discussed. PMID:25129742

Mushi, Ngesa Ezekiel; Butchosa, Nuria; Salajkova, Michaela; Zhou, Qi; Berglund, Lars A

2014-11-01

269

Characterization and cloning of chitin deacetylases from Rhizopus circinans.  

PubMed

Chitin deacetylase catalyzes hydrolysis of the acetamido groups of N-acetylglucosamine of chitin in fungal cell walls. Here a chitin deacetylase secreted by Rhizopus circinans was purified to homogeneity and partially characterized. The enzyme exhibits an apparent molecular weight of approximately 75kDa. At 37 degrees C it shows optimal activity at pH 5.5-6. Its pH stability and thermal stability are good. Mn(2+) and Mg(2+) slightly enhance the activity of the enzyme and Cu(2+) strongly inhibits it. An R. circinans cDNA library was constructed and screened with a homologous probe synthesized by RT-PCR or with synthetic primers derived from the N-terminal amino-acid sequence of the native purified chitin deacetylase. Three chitin deacetylase cDNAs (RC, D2, and I3/2) were isolated from the cDNA library and sequenced. These cDNAs exhibit features characteristic of chitin deacetylase sequences: the presence of a polysaccharide deacetylase domain, a metal-binding triad, the conserved catalytic residues, and high homology with various chitin deacetylase genes. The cDNAs were cloned in a Pichia pastoris expression system and produced as polyhistidine-tagged proteins. Only one recombinant enzyme (called RC) was active under the tested conditions. It was purified to homogeneity in a single step and further characterized. The protein showed an apparent molecular mass of approximately 75kDa and, like the native enzyme, showed optimal activity at pH 5.5-6 at 37 degrees C. It was strongly inhibited by Cu(2+). The isolation of several chitin deacetylase cDNAs from the same microorganism is discussed. PMID:18314348

Gauthier, Carole; Clerisse, Fabienne; Dommes, Jacques; Jaspar-Versali, Marie-France

2008-05-01

270

Chitin-microparticles for the control of intestinal inflammation  

PubMed Central

Chitin is a polymer of N-acetylglucosamine with the ability to regulate innate and adaptive immune responses. However, the detailed mechanisms of chitin-mediated regulation of intestinal inflammation are only partially known. In this study, Chitin-microparticles (CMPs) or PBS were orally administered to acute and chronic colitis models every three days for six consecutive weeks beginning at weaning age. The effects of this treatment were evaluated by histology, cytokine production, co-culture study and enteric bacterial analysis in DSS-induced colitis or TCR? knockout chronic colitis models. Histologically, chitin-treated mice showed significantly suppressed colitis as compared to PBS-treated mice in both animal models. The production of IFN? was upregulated in the mucosa of chitin-treated mice compared to control mice. The major source of IFN?-producing cells was CD4+ T cells. In mouse dendritic cells (DCs), we found that CMPs were efficiently internalized and processed within 48 hours. Mesenteric lymph nodes (MLNs) CD4+ T cells isolated from chitin-treated mice produced 7-fold higher amount of IFN? in the culture supernatant after being co-cultured with DCs and chitin as compared to the control. Proliferation of CFSElow CD4+ T cells in MLNs and enteric bacterial translocation rates were significantly reduced in chitin-treated mice when compared to the control. In addition, CMPs improved the imbalance of enteric bacterial compositions and significantly increased IL-10-producing cells in non-inflamed colon, indicating the immunoregulatory effects of CMPs in intestinal mucosa. In conclusion, CMPs significantly suppress the development of inflammation by modulating cytokine balance and microbial environment in colon. PMID:22241684

Nagatani, Katsuya; Wang, Sen; Llado, Victoria; Lau, Cindy W.; Li, Zongxi; Mizoguchi, Atsushi; Nagler, Cathryn R.; Shibata, Yoshimi; Reinecker, Hans-Christian; Mora, J. Rodrigo; Mizoguchi, Emiko

2013-01-01

271

Growth of calcium phosphate on phosphorylated chitin fibres  

Microsoft Academic Search

Calcium phosphate growth on chitin phosphorylated fibres was studied using scanning electron microscopy and energy dispersive X-ray analysis (SEM, EDX), micro-Fourier transform infrared spectroscopy (FTIR), and solid state magic angle spinning nuclear magnetic resonance (MAS NMR) techniques. The C6 chemical shift positions of 13C MAS NMR in the chitin fibres phosphorylated using urea and H3PO4 are obvious indicating that phosphorylation

Y. YOKOGAWA; J PAZ REYES; M. R MUCALO; M TORIYAMA; Y KAWAMOTO; T SUZUKI; K NISHIZAWA; F NAGATA; T KAMAYAMA

1997-01-01

272

Digestibility of chitin in cod, Gadus morhua, in vivo  

NASA Astrophysics Data System (ADS)

Sixteen cod, Gadus morhua (L.), were individually fed a single ration of shrimps, Crangon allmanni. Four fish were killed and examined 6, 12, 24 and 48 h after the fish had been fed. Chitinase activities were measured in the extracts of stomach contents, stomach tissue, pyloric caecae, intestinal contents and intestinal tissue. The level of enzyme activity in different parts of the digestive tract was shown to be dependent on the phase of the digestive process. High concentrations of the chitin degradation product N-acetyl-D-glucosamine were determined in the stomach and in the intestinal contents. Based on the chitin concentration in the food organisms and the individual food uptake, the amount of chitin consumed by each fish could be calculated. Only up to 9% of the ingested chitin was recovered from the intestinal contents of the fish at any given time after feeding (6, 12, 24 and 48 h). In addition, only 2.4% of the chitin consumed with the food could be recovered in the collected faeces of the fish. The 4 cod killed 48 h after feeding had completely emptied their stomach. Chitin digestion in these fish was calculated to have been 90%.

Danulat, Eva

1987-12-01

273

Chitin membranes containing silver nanoparticles for wound dressing application.  

PubMed

Silver nanoparticles are gaining importance as an antimicrobial agent in wound dressings. Chitin is a biopolymer envisioned to promote rapid dermal regeneration and accelerate wound healing. This study was focused on the evaluation of chitin membranes containing silver nanoparticles for use as an antimicrobial wound dressing. Silver nanoparticles were synthesised by gamma irradiation at doses of 50 kGy in the presence of sodium alginate as stabiliser. The UV-Vis absorption spectra of nanoparticles exhibited an absorption band at 415-420 nm, which is the typical plasmon resonance band of silver nanoparticles. The peaks in the X-ray diffraction (XRD) pattern are in agreement with the standard values of the face-centred cubic silver. Transmission electron microscopy (TEM) images indicate silver nanoparticles with spherical morphology and small particle size in the range of 3-13 nm. In vitro antimicrobial tests were performed using Pseudomonas aeruginosa and Staphylococcus aureus to determine the antimicrobial efficiency of the chitin membranes containing 30, 50, 70 and 100 ppm nanosilver. No viable counts for P. aeruginosa were detected with 70 ppm silver nanoparticles dressing after 1-hour exposure. A 2-log reduction in viable cell count was observed for S. aureus after 1 hour and a 4-log reduction after 6 hours with 100 ppm nanosilver chitin membranes. This study demonstrates the antimicrobial capability of chitin membranes containing silver nanoparticles. The chitin membranes with 100 ppm nanosilver showed promising antimicrobial activity against common wound pathogens. PMID:22958740

Singh, Rita; Singh, Durgeshwer

2014-06-01

274

Bacterial Chitin Hydrolysis in Two Lakes with Contrasting Trophic Statuses  

PubMed Central

Chitin, which is a biopolymer of the amino sugar glucosamine (GlcN), is highly abundant in aquatic ecosystems, and its degradation is assigned a key role in the recycling of carbon and nitrogen. In order to study the significance of chitin decomposition in two temperate freshwater lakes with contrasting trophic and redox conditions, we measured the turnover rate of the chitin analog methylumbelliferyl-N,N?-diacetylchitobioside (MUF-DC) and the presence of chitinase (chiA) genes in zooplankton, water, and sediment samples. In contrast to the eutrophic and partially anoxic lake, chiA gene fragments were detectable throughout the oligotrophic water column and chiA copy numbers per ml of water were up to 15 times higher than in the eutrophic waters. For both lakes, the highest chiA abundance was found in the euphotic zone—the main habitat of zooplankton, but also the site of production of easily degradable algal chitin. The bulk of chitinase activity was measured in zooplankton samples and the sediments, where recalcitrant chitin is deposited. Both, chiA abundance and chitinase activity correlated well with organic carbon, nitrogen, and concentrations of particulate GlcN. Our findings show that chitin, although its overall contribution to the total organic carbon is small (?0.01 to 0.1%), constitutes an important microbial growth substrate in these temperate freshwater lakes, particularly where other easily degradable carbon sources are scarce. PMID:22101058

Carstens, Dörte; Keller, Esther; Vazquez, Francisco; Schubert, Carsten J.; Zeyer, Josef; Bürgmann, Helmut

2012-01-01

275

In vivo hyaluronan synthesis upon expression of the mammalian hyaluronan synthase gene in Drosophila.  

PubMed

Hyaluronan (HA) is a large linear polymer of repeating disaccharides of glucuronic acid and GlcNAc. Although HA is widely distributed in vertebrate animals, it has not been found in invertebrates, including insect species. Insects utilize chitin, a repeating beta-1,4-linked homopolymer of GlcNAc, as a major component of their exoskeleton. Recent studies illustrate the similarities in the biosynthetic mechanisms of HA and chitin and suggest that HA synthase (HAS) and chitin synthase have evolved from a common ancestral molecule. Although the biochemical properties and in vivo functions of HAS proteins have been extensively studied, the molecular basis for HA biosynthesis is not completely understood. For example, it is currently not clear if proper chain elongation and secretion of HA require other components in addition to HAS. Here, we demonstrate that a non-HA-synthesizing animal, the fruit fly Drosophila melanogaster, can produce HA in vivo when a single HAS protein is introduced. Expression of the mouse HAS2 gene in Drosophila tissues by the Gal4/UAS (upstream activating sequence) system resulted in massive HA accumulation in the extracellular space and caused various morphological defects. These morphological abnormalities were ascribed to disordered cell-cell communications due to accumulation of HA rather than disruption of heparan sulfate synthesis. We also show that adult wings with HA can hold a high level of water. These findings demonstrate that organisms synthesizing chitin (but not HA) are capable of producing HA that is structurally and functionally relevant to that in mammals. The ability of insect cells to produce HA supports the idea that in vivo HA biosynthesis does not require molecules other than the HAS protein. An alternative model is that Drosophila cells use endogenous components of the chitin biosynthetic machinery to produce and secrete HA. PMID:14966127

Takeo, Satomi; Fujise, Momoko; Akiyama, Takuya; Habuchi, Hiroko; Itano, Naoki; Matsuo, Takashi; Aigaki, Toshiro; Kimata, Koji; Nakato, Hiroshi

2004-04-30

276

Structural differences between chitin and chitosan extracted from three different marine sources.  

PubMed

Three marine sources of chitin from Tunisia were investigated. Structural differences between ?-chitin from shrimp (Penaeus kerathurus) waste, crab (Carcinus mediterraneus) shells, and ?-chitin from cuttlefish (Sepia officinalis) bones were studied by the (13)C NMR, FTIR, and XRD diffractograms. The (13)C NMR analysis showed a splitting of the C3 and C5 carbon signals for ?-chitin, while that of ?-chitin was merged into a single resonance. The bands contour of deconvoluted and curve-fit FTIR spectra showed a more detailed structure of ?-chitin in the region of O-H, N-H and CO stretching regions. IR and (13)C NMR were used to determine the chitin degree of acetylation (DA). XRD analysis indicated that ?-chitins were more crystalline polymorph than ?-chitin. Shrimp chitin was obtained with a good yield (20% on raw material dry weight) and no residual protein and salts. Chitosans, with a DA lower than 20% and relatively low molecular masses were prepared from the wet chitins in the same experimental conditions. They were perfectly soluble in acidic medium. Nevertheless, chitin and chitosan characteristics were depending upon the chitin source. PMID:24468048

Hajji, Sawssen; Younes, Islem; Ghorbel-Bellaaj, Olfa; Hajji, Rachid; Rinaudo, Marguerite; Nasri, Moncef; Jellouli, Kemel

2014-04-01

277

XRD studies of chitin-based polyurethane elastomers.  

PubMed

Chitin-based polyurethane elastomers (PUEs) were synthesized by step growth polymerization techniques using poly(epsilon-caprolactone) (PCL) varying diisocyanate and chain extender structures. The viscosity average molecular weight (M(v)) of chitin was deduced from the intrinsic viscosity and found; M(v)=6.067 x 10(5). The conventional spectroscopic characterization of the samples with FTIR, (1)H NMR and (13)C NMR were in accordance with proposed PUEs structure. The crystalline behavior of the synthesized polymers were investigated by X-ray diffraction (XRD), differential scanning calorimetery (DSC) and loss tangent curves (tan delta peaks). The observed patterns of the crystalline peaks for the lower angle for chitin in the 2theta range were indexed as 9.39 degrees, 19.72 degrees, 20.73 degrees, 23.41 degrees and 26.39 degrees. Results showed that crystallinity of the synthesized PUEs samples was affected by varying the structure of the diisocyanate and chain extender. Crystallinity decreased from aliphatic to aromatic characters of the diisocyanates used in the final PU. The presence of chitin also favors the formation of more ordered structure, as higher peak intensities was obtained from the PU extended with chitin than 1,4-butane diol (BDO). The value of peak enthalpy (DeltaH) of chitin was found to be 47.13 J g(-1). The higher DeltaH value of 46.35 J g(-1) was found in the samples extended with chitin than BDO (39.73 J g(-1)). PMID:18495239

Zia, Khalid Mahmood; Bhatti, Ijaz Ahmad; Barikani, Mehdi; Zuber, Mohammad; Sheikh, Munir Ahmad

2008-08-15

278

Chitin hydrolysis by Listeria spp., including L. monocytogenes.  

PubMed

Listeria spp., including the food-borne pathogen Listeria monocytogenes, are ubiquitous microorganisms in the environment and thus are difficult to exclude from food processing plants. The factors that contribute to their multiplication and survival in nature are not well understood, but the ability to catabolize various carbohydrates is likely to be very important. One major source of carbon and nitrogen in nature is chitin, an insoluble linear beta-1,4-linked polymer of N-acetylglucosamine (GlcNAc). Chitin is found in cell walls of fungi and certain algae, in the cuticles of arthropods, and in shells and radulae of molluscs. In the present study, we demonstrated that L. monocytogenes and other Listeria spp. are able to hydrolyze alpha-chitin. The chitinolytic activity is repressed by the presence of glucose in the medium, suggesting that chitinolytic activity is subjected to catabolite repression. Activity is also regulated by temperature and is higher at 30 degrees C than at 37 degrees C. In L. monocytogenes EGD, chitin hydrolysis depends on genes encoding two chitinases, lmo0105 (chiB) and lmo1883 (chiA), but not on a gene encoding a putative chitin binding protein (lmo2467). The chiB and chiA genes are phylogenetically related to various well-characterized chitinases. The potential biological implications of chitinolytic activity of Listeria are discussed. PMID:18424542

Leisner, J J; Larsen, M H; Jørgensen, R L; Brøndsted, L; Thomsen, L E; Ingmer, H

2008-06-01

279

Chitin in the silk gland ducts of the spider Nephila edulis and the silkworm Bombyx mori.  

PubMed

Here we report the detection and localisation of chitin in the cuticle of the spinning ducts of both the spider Nephila edulis and the silkworm Bombyx mori. Our observations demonstrate that the duct walls of both animals contain chitin notwithstanding totally independent evolutionary pathways of the systems. We conclude that chitin may well be an essential component for the construction of spinning ducts; we further conclude that in both species chitin may indicate the evolutionary origin of the spinning ducts. PMID:24015298

Davies, Gwilym J G; Knight, David P; Vollrath, Fritz

2013-01-01

280

YEA! ® Elicitor Response Comparison to Chitin \\/ Chitosan in Mung Bean and Adzuki Bean Germination Experiments  

Microsoft Academic Search

YEA! ® (1 mg\\/seed) to the 0.9 mg\\/seed treatment with chitin \\/ chitosan elicited five times as much 9-1,3-glucanase enzyme activity. Secondly, lower concentrations of the chitin oligosaccharide containing six glycan moieties, N-acetylchitohexaose were studied. The importance of the chitin oligosaccharide is that short chains of chitin have been found optimal in elicitation of many types of plants. The dose

James C. Linden; Richard J. Stoner

281

Effects of molecular weight and deacetylation degree of chitin\\/chitosan on wound healing  

Microsoft Academic Search

We studied the effects of chitin\\/chitosan on wound healing with reference to chemical properties using a linear incisional wound model in rats. Wound break strength of the chitosan group (D-glucosamine (GlcN), chito-oligosaccharide (COS), chitosan) was higher than the chitin group (N-acetyl-D-glucosamine (GlcNAc), chiti-oligosaccharide (NACOS), chitin). Collagenase activity was also higher in the chitosan group than the chitin group. There was

Tatsuya Minagawa; Yasuhiko Okamura; Yoshihiro Shigemasa; Saburo Minami; Yoshiharu Okamoto

2007-01-01

282

Biomimetic nanofibrous scaffolds: Preparation and characterization of chitin\\/silk fibroin blend nanofibers  

Microsoft Academic Search

Electrospinning of chitin\\/silk fibroin (SF) blend solutions in 1,1,1,3,3,3-hexafluoro-2-propanol (HFIP) was investigated to fabricate a biomimetic nanostructured scaffolds for tissue engineering. The morphology of the electrospun chitin\\/SF blend nanofibers was investigated with a field emission scanning electron microscope (FE-SEM). The average diameters of chitin\\/SF blend fibers decreased from 920 to 340nm, with the increase of chitin content in blend compositions.

Ko Eun Park; Sung Youn Jung; Seung Jin Lee; Byung-Moo Min; Won Ho Park

2006-01-01

283

Chitin in the Silk Gland Ducts of the Spider Nephila edulis and the Silkworm Bombyx mori  

PubMed Central

Here we report the detection and localisation of chitin in the cuticle of the spinning ducts of both the spider Nephila edulis and the silkworm Bombyx mori. Our observations demonstrate that the duct walls of both animals contain chitin notwithstanding totally independent evolutionary pathways of the systems. We conclude that chitin may well be an essential component for the construction of spinning ducts; we further conclude that in both species chitin may indicate the evolutionary origin of the spinning ducts. PMID:24015298

Davies, Gwilym J. G.; Knight, David P.; Vollrath, Fritz

2013-01-01

284

The biocompatibility of dibutyryl chitin in the context of wound dressings  

Microsoft Academic Search

Dibutyryl chitin (DBC) is a modified chitin carrying butyryl groups at 3 and 6 positions; its peculiarity is that it dissolves promptly in common solvents, while being insoluble in aqueous systems. The high biocompatibility of dibutyryl chitin in the form of films and non-wovens has been demonstrated for human, chick and mouse fibroblasts by the Viability\\/Cytotoxicity assay, In situ Cell

Riccardo A. A. Muzzarelli; Mario Guerrieri; Gaia Goteri; Corrado Muzzarelli; Tatiana Armeni; Roberto Ghiselli; Maria Cornelissen

2005-01-01

285

Comparative protein analysis of the chitin metabolic pathway in extant organisms: A complex network approach  

Microsoft Academic Search

Chitin is a structural endogenous carbohydrate, which is a major component of fungal cell walls and arthropod exoskeletons. A renewable resource and the second most abundant polysaccharide in nature after cellulose, chitin is currently used for waste water clearing, cosmetics, medical, and veterinary applications. This work comprises data mining of protein sequences related to the chitin metabolic pathway of completely

Aristoteles Goes-Neto; Marcelo V. C. Diniz; Leonardo B. L. Santos; Suani T. R. Pinho; José G. V. Miranda; Thierry Petit Lobao; Ernesto P. Borges; Charbel Niño El-Hani; Roberto F. S. Andrade

2010-01-01

286

Antipsychotic, antidepressant, anxiolytic, and anticonvulsant drugs induce type II nitric oxide synthase mRNA in rat brain  

Microsoft Academic Search

Nitric oxide synthase inhibitors have been regarded as potentially beneficial for psychiatric disorders such as depression and schizophrenia, though little is known about how nitric oxide synthases are affected by psychotropic drugs in the brain. Using reverse transcription-polymerase chain reaction analysis, we investigated the effects of short- and long-term oral treatments with several psychotropics on type II nitric oxide synthase

Eiji Suzuki; Toshio Nakaki; Futoshi Shintani; Shigenobu Kanba; Hitoshi Miyaoka

2002-01-01

287

Ultrasonication and steam-explosion as chitin pretreatments for chitin oligosaccharide production by chitinases of Lecanicillium lecanii.  

PubMed

In this study, chitin oligosaccharides have been successfully produced using chitinases from submerged fermentation of Lecanicillium lecanii. The highest Hex, Chit and Prot production was 0.14, 0.26 and 2.05 U/mg of protein, respectively, which were attained varying pH from 5 to 8 after 96 h. Culture conditions conducted at constant pH of 6 resulted in significantly lower enzyme production. The crude enzyme was partially purified by salting out with (NH4)2SO4 followed by size exclusion chromatography to isolate the chitinase mixture for further chitin hydrolysis assays. In this regard, chitin substrates were pretreated with sonication and steam explosion prior to enzymatic reaction. Structural changes were observed with steam explosion with 11.28% reduction of the crystallinity index attained with the lowest chitin/water ratio (0.1g/mL). Pretreated chitins reached the highest production of reducing sugars (0.37 mg/mL) and GlcNAc (0.59 mg/mL) in 23.6% yield. PMID:23993287

Villa-Lerma, Guadalupe; González-Márquez, Humberto; Gimeno, Miquel; López-Luna, Alberto; Bárzana, Eduardo; Shirai, Keiko

2013-10-01

288

Polarized light-stimulated enzymatic hydrolysis of chitin and chitosan.  

PubMed

Illumination with white linearly polarized light (WLPL) stimulated chitinase and chitosanase in their degradation of chitin and chitosan, respectively. Enzymes were illuminated at room temperature in separate vessels, then admixed in reactors containing polysaccharides. Hydrolysis of chitosan to glucosamine followed first order kinetics whereas hydrolysis of chitin to N-acetylglucosamine deviated from the first order kinetics. In both cases, an increase in the rate of hydrolysis depended on the illumination time. Efficient degradation required up to 60 min exposure of the enzyme to WLPL. PMID:18823881

Konieczna-Molenda, Anna; Fiedorowicz, Maciej; Zhong, Wei; Tomasik, Piotr

2008-12-01

289

INHIBITOR STUDIES ON MYCOBACTERIUM TUBERCULOSIS MALATE SYNTHASE  

E-print Network

to conventional drug therapies, is of particular interest. Persistent bacteria rely on metabolic pathways that are distinct from active infection Mtb as the environmental conditions of the persistent state are different (e.g., low nutrient). Because persistent...

Owen, Joshua

2008-08-03

290

Nitric Oxide Synthase Inhibitors as Antidepressants  

PubMed Central

Affective and anxiety disorders are widely distributed disorders with severe social and economic effects. Evidence is emphatic that effective treatment helps to restore function and quality of life. Due to the action of most modern antidepressant drugs, serotonergic mechanisms have traditionally been suggested to play major roles in the pathophysiology of mood and stress-related disorders. However, a few clinical and several pre-clinical studies, strongly suggest involvement of the nitric oxide (NO) signaling pathway in these disorders. Moreover, several of the conventional neurotransmitters, including serotonin, glutamate and GABA, are intimately regulated by NO, and distinct classes of antidepressants have been found to modulate the hippocampal NO level in vivo. The NO system is therefore a potential target for antidepressant and anxiolytic drug action in acute therapy as well as in prophylaxis. This paper reviews the effect of drugs modulating NO synthesis in anxiety and depression.

Wegener, Gregers; Volke, Vallo

2010-01-01

291

Mechanism of Action and Inhibition of dehydrosqualene Synthase  

Microsoft Academic Search

'Head-to-head' terpene synthases catalyze the first committed steps in sterol and carotenoid biosynthesis: the condensation of two isoprenoid diphosphates to form cyclopropylcarbinyl diphosphates, followed by ring opening. Here, we report the structures of Staphylococcus aureus dehydrosqualene synthase (CrtM) complexed with its reaction intermediate, presqualene diphosphate (PSPP), the dehydrosqualene (DHS) product, as well as a series of inhibitors. The results indicate

F Lin; C Liu; Y Liu; Y Zhang; K Wang; W Jeng; T Ko; R Cao; A Wang; E Oldfield

2011-01-01

292

Cellulose synthase interacting protein  

PubMed Central

Cellulose is the most abundant biopolymer on earth. The great abundance of cellulose places it at the forefront as a primary source of biomass for renewable biofuels. However, the knowledge of how plant cells make cellulose remains very rudimentary. Cellulose microfibrils are synthesized at the plasma membrane by hexameric protein complexes, also known as cellulose synthase complexes. The only known components of cellulose synthase complexes are cellulose synthase (CESA) proteins until the recent identification of a novel component. CSI1, which encodes CESA interacting protein 1 (CSI1) in Arabidopsis. CSI1, as the first non-CESA proteins associated with cellulose synthase complexes, opens up many opportunities. PMID:21150290

Somerville, Chris

2010-01-01

293

Preparation of chitin-silica composites by in vitro silicification of two-dimensional Ianthella basta demosponge chitinous scaffolds under modified Stöber conditions.  

PubMed

Chitin is a biopolymer found in cell walls of various fungi and skeletal structures of numerous invertebrates. The occurrence of chitin within calcium- and silica-containing biominerals has inspired development of chitin-based hybrids and composites in vitro with specific physico-chemical and material properties. We show here for the first time that the two-dimensional ?-chitin scaffolds isolated from the skeletons of marine demosponge Ianthella basta can be effectively silicified by the two-step method with the use of Stöber silica micro- and nanodispersions under Extreme Biomimetic conditions. The chitin-silica composites obtained at 120 °C were characterized by the presence of spherical SiO2 particles homogeneously distributed over the chitin fibers, which probably follows from the compatibility of Si-OH groups to the hydroxyl groups of chitin. The biocomposites obtained were characterized by various analytical techniques such as energy dispersive spectrometry, scanning electron microscopy, thermogravimetric/differential thermal analyses as well as X-ray photoelectron spectroscopy, Fourier transform infrared and Raman spectroscopy to determine possible interactions between silica and chitin molecule. The results presented proved that the character and course of the in vitro chitin silicification in Stöber dispersions depended considerably on the degree of hydrolysis of the SiO2 precursor. PMID:23910299

Wysokowski, Marcin; Behm, Thomas; Born, René; Bazhenov, Vasilii V; Meissner, Heike; Richter, Gert; Szwarc-Rzepka, Karolina; Makarova, Anna; Vyalikh, Denis; Schupp, Peter; Jesionowski, Teofil; Ehrlich, Hermann

2013-10-01

294

Development and Binding Mode Assessment of N-[4-[2-propyn-1-yl[(6S)-4,6,7,8-tetrahydro-2-(hydroxymethyl)-4-oxo-3H-cyclopenta[g]quinazolin-6-yl]amino]benzoyl]-L-?-glutamyl-D-glutamic acid (BGC 945), a Novel Thymidylate Synthase Inhibitor that Targets Tumor Cells  

PubMed Central

N-[4-[2-propyn-1-yl[(6S)-4,6,7,8-tetrahydro-2-(hydroxymethyl)-4-oxo-3H-cyclopenta[g]quinazolin-6-yl]amino]benzoyl]-L-?-glutamyl-D-glutamic acid 1 (BGC 945, now known as ONX 0801), is a small molecule thymidylate synthase (TS) inhibitor discovered at the Institute of Cancer Research in London. It is licensed by Onyx Pharmaceuticals and is in Phase 1 clinical studies. It is a novel antifolate drug resembling TS inhibitors plevitrexed and raltitrexed that combines enzymatic inhibition of thymidylate synthase with ?-folate receptor-mediated targeting of tumor cells. Thus, it has potential for efficacy with lower toxicity due to selective intracellular accumulation through ?-folate receptor (?-FR) transport. The ?-FR, a cell-surface receptor glycoprotein, which is over expressed mainly in ovarian and lung cancer tumors, has an affinity for 1 similar to that for its natural ligand, folic acid. This study describes a novel synthesis of 1, an X-ray crystal structure of its complex with Escherichia coli TS and 2’-deoxyuridine-5’-monophosphate, and a model for a similar complex with human TS. PMID:23710599

Tochowicz, Anna; Dalziel, Sean; Eidam, Oliv; O’Connell, Joseph D.; Griner, Sarah; Finer-Moore, Janet S.; Stroud, Robert M.

2013-01-01

295

Characterization of Chitin and Chitosan Molecular Structure in Aqueous Solution  

SciTech Connect

Molecular dynamics simulations have been used to characterize the structure of chitin and chitosan fibers in aqueous solutions. Chitin fibers, whether isolated or in the form of a ?-chitin nanoparticle, adopt the so-called 2-fold helix with ? and ? values similar to its crystalline state. In solution, the intramolecular hydrogen bond HO3(n)?O5(n+1) responsible for the 2-fold helical motif is stabilized by hydrogen bonds with water molecules in a well-defined orientation. On the other hand, chitosan can adopt five distinct helical motifs and its conformational equilibrium is highly dependent on pH. The hydrogen bond pattern and solvation around the O3 atom of insoluble chitosan (basic pH) are nearly identical to these quantities in chitin. Our findings suggest that the solubility and conformation of these polysaccharides are related to the stability of the intrachain HO3(n)?O5(n+1) hydrogen bond, which is affect by the water exchange around the O3-HO3 hydroxyl group.

Franca, Eduardo D.; Lins, Roberto D.; Freitas, Luiz C.; Straatsma, TP

2008-12-01

296

Physiology of microbial degradation of chitin and chitosan  

Microsoft Academic Search

Chitin is produced in enormous quantities in the biosphere, chiefly as the major structural component of most fungi and invertebrates. Its degradation is chiefly by bacteria and fungi, by chitinolysis via chitinases, but also via deacetylation to chitosan, which is hydrolysed by chitosanases. Chitinases and chitosanases have a range of roles in the organisms producing them: autolytic, morphogenetic or nutritional.

Graham W. Gooday

1990-01-01

297

Growth of calcium phosphate on phosphorylated chitin fibres.  

PubMed

Calcium phosphate growth on chitin phosphorylated fibres was studied using scanning electron microscopy and energy dispersive X-ray analysis (SEM, EDX), micro-Fourier transform infrared spectroscopy (FTIR), and solid state magic angle spinning nuclear magnetic resonance (MAS NMR) techniques. The C6 chemical shift positions of 13C MAS NMR in the chitin fibres phosphorylated using urea and H3PO4 are obvious indicating that phosphorylation takes place not in the C1 but in the C6 region. Micro-FTIR and 31P MAS NMR suggested that ammonium hydrogen phosphate formed during the phosphorylation procedure. Chitin fibres phosphorylated using urea and H3PO4 and then soaked in saturated Ca(OH)2 solution at ambient temperature, which lead to the formation of thin coatings formed by partial hydrolysis of the PO4 functionalities, were found to stimulate the growth of a calcium phosphate coating on their surfaces after soaking in 1.5xSBF solution for as little as one day. The thin layer after Ca(OH)2 treatment functioned as a nucleation layer for further calcium phosphate deposition after soaking in 1.5xSBF solution. EDX-measured Ca : P ratios of the coatings of Ca(OH)2-treated phosphorylated chitin in 1.5xSBF solution suggested that calcium-deficient apatite was formed. PMID:15348722

Yokogawa, Y; Paz Reyes, J; Mucalo, M R; Toriyama, M; Kawamoto, Y; Suzuki, T; Nishizawa, K; Nagata, F; Kamayama, T

1997-07-01

298

Dynamics of Gram-negative bacteria population density in a soil in the course of the succession initiated by chitin and cellulose  

NASA Astrophysics Data System (ADS)

The functions of actinomycetes in polymer destruction in soil traditionally considered as the dominant, compare to another groups of bacteria. Gram-positive bacteria also have ecological functions in destruction of soil organic matter. The role of Gram-negative bacteria has been researched in the microbial succession in terms of polymers destruction, which are widely spreads in soils: chitin and cellulose. The method with nalidixic acid as an inhibitor of DNA division of Gram-negative bacteria was modified. By modified method microbial succession of Gram-negative bacteria in the different horizons of a chernozem under aerobic and anaerobic conditions was researched. Chitin and cellulose as the source of nutrients with moistening was used in experiments. The introduction of chitin had no positive effect on the population density of Gram-negative bacteria in a chernozem, but it advanced the date of their appearance in microbial succession: the maximum of Gram-negative bacteria population density was registered on the 3rd- 7th day of the experiment with adding chitin. Compare to the control, which one was without any nutrient adding this dynamics registered much earlier. Consequently, the introduction of chitin as an additional source of nutrition promoted revealing of the Gram-negative bacteria in soil already at the early stages of the succession. In the course of the succession, when the fungal mycelium begins to die off, the actinomycetic mycelium increases in length, i.e., Gram-negative bacteria are replaced at this stage with Gram-positive ones, the leading role among which belongs to actinomycetes. The growth rate of Gram-negative bacteria is higher than that of actinomycetes, so they start chitin utilization at the early stages of the succession, whereas actinomycetes dominate at the late stages. The population density of Gram-negative bacteria was lower under the anaerobic conditions as compared with that in the aerobic ones. The population density of Gram-negative bacteria in the lower layer of the A horizon of the chernozem and in the B horizon was slightly higher only in the case of the chitin introduction. When cellulose was introduced into the soil under aerobic conditions, the population density of Gram-negative bacteria in all the layers of the A horizon of the chernozem was maximal from the 14th to the 22nd day of the experiment. Simultaneously, an increase in the length of the actinomycetal mycelium was observed, as these organisms also perform cellulose hydrolysis in soils. The Gram-negative bacteria began to develop at the stage of the fungal mycelium destruction, which indirectly confirmed the chitinolytic activity of these bacteria.

Konstantin, Ivanov; Lubov, Polyanskaya

2014-05-01

299

Nikkomycin Z is an effective inhibitor of the chytrid fungus linked to global amphibian declines.  

PubMed

Fungal infections in humans, wildlife, and plants are a growing concern because of their devastating effects on human and ecosystem health. In recent years, populations of many amphibian species have declined, and some have become extinct due to chytridiomycosis caused by the fungal pathogen Batrachochytrium dendrobatidis. For some endangered amphibian species, captive colonies are the best intermediate solution towards eventual reintroduction, and effective antifungal treatments are needed to cure chytridiomycosis and limit the spread of this pathogen in such survival assurance colonies. Currently, the best accepted treatment for infected amphibians is itraconazole, but its toxic side effects reduce its usefulness for many species. Safer antifungal treatments are needed for disease control. Here, we show that nikkomycin Z, a chitin synthase inhibitor, dramatically alters the cell wall stability of B. dendrobatidis cells and completely inhibits growth of B. dendrobatidis at 250 ?M. Low doses of nikkomycin Z enhanced the effectiveness of natural antimicrobial skin peptide mixtures tested in vitro. These studies suggest that nikkomycin Z would be an effective treatment to significantly reduce the fungal burden in frogs infected by B. dendrobatidis. PMID:24433676

Holden, Whitney M; Fites, J Scott; Reinert, Laura K; Rollins-Smith, Louise A

2014-01-01

300

Customizing Properties of ?-Chitin in Squid Pen (Gladius) by Chemical Treatments.  

PubMed

The squid pen (gladius) from the Loligo vulgaris was used for preparation of ?-chitin materials characterized by different chemical, micro- and nano-structural properties that preserved, almost completely the macrostructural and the mechanical ones. The ?-chitin materials obtained by alkaline treatment showed porosity, wettability and swelling that are a function of the duration of the treatment. Microscopic, spectroscopic and synchrotron X-ray diffraction techniques showed that the chemical environment of the N-acetyl groups of the ?-chitin chains changes after the thermal alkaline treatment. As a consequence, the crystalline packing of the ?-chitin is modified, due to the intercalation of water molecules between ?-chitin sheets. Potential applications of these ?-chitin materials range from the nanotechnology to the regenerative medicine. The use of gladii, which are waste products of the fishing industry, has also important environmental implications. PMID:25517216

Ianiro, Alessandro; Giosia, Matteo Di; Fermani, Simona; Samorì, Chiara; Barbalinardo, Marianna; Valle, Francesco; Pellegrini, Graziella; Biscarini, Fabio; Zerbetto, Francesco; Calvaresi, Matteo; Falini, Giuseppe

2014-12-01

301

Customizing Properties of ?-Chitin in Squid Pen (Gladius) by Chemical Treatments  

PubMed Central

The squid pen (gladius) from the Loligo vulgaris was used for preparation of ?-chitin materials characterized by different chemical, micro- and nano-structural properties that preserved, almost completely the macrostructural and the mechanical ones. The ?-chitin materials obtained by alkaline treatment showed porosity, wettability and swelling that are a function of the duration of the treatment. Microscopic, spectroscopic and synchrotron X-ray diffraction techniques showed that the chemical environment of the N-acetyl groups of the ?-chitin chains changes after the thermal alkaline treatment. As a consequence, the crystalline packing of the ?-chitin is modified, due to the intercalation of water molecules between ?-chitin sheets. Potential applications of these ?-chitin materials range from the nanotechnology to the regenerative medicine. The use of gladii, which are waste products of the fishing industry, has also important environmental implications. PMID:25517216

Ianiro, Alessandro; Di Giosia, Matteo; Fermani, Simona; Samorì, Chiara; Barbalinardo, Marianna; Valle, Francesco; Pellegrini, Graziella; Biscarini, Fabio; Zerbetto, Francesco; Calvaresi, Matteo; Falini, Giuseppe

2014-01-01

302

Effect of soluble polysaccharides addition on rheological properties and microstructure of chitin nanocrystal aqueous dispersions.  

PubMed

Mixtures of chitin nanocrystal aqueous dispersions (at pH 3.0) with soluble polysaccharides of varying molecular features were examined rheologically and microscopically, under different conditions of biopolymer concentration, ionic strength, pH and temperature. The addition of non-adsorbing polysaccharides (guar gum, locust bean gum and xanthan) as well as oppositely charged (?-carrageenan) to a chitin nanocrystal dispersion, resulted in a network formation and the gel strength increased with the chitin nanocrystal concentration. In contrast, the chitin nanocrystal - chitosan or - pullulan mixed dispersions did not show any network formation (tan?>1) at the concentration range examined. An increase in ionic strength and pH also resulted in an enhanced elasticity of the chitin nanocrystal-guar gum dispersions. Furthermore, an increase in the elastic modulus, which was irreversible upon cooling, was observed upon heating the chitin nanocrystal-polysaccharide mixed dispersions. PMID:23618276

Tzoumaki, Maria V; Moschakis, Thomas; Biliaderis, Costas G

2013-06-01

303

Functionality of chitin as a direct compression excipient: an acetaminophen comparative study.  

PubMed

The particle and tableting properties of chitin extracted from shrimp exoskeletons were evaluated and compared with common excipients used for the preparation of tablets. Chitin offered more benefits in terms of functionality than calcium diphosphate, lactose monohydrate and pregelatinized starch. Further, highly plastic deforming materials such as sorbitol and PVP K30 and microcrystalline cellulose showed the best compactibility and dilution potential, whereas brittle deforming materials such as lactose monohydrate and calcium diphosphate were poorly compactable. Chitin had better compactibility than pregelatinized starch, calcium diphosphate and lactose monohydrate. Further, along with calcium diphosphate, chitin was the least sensitive material to compaction speed due to a combination of a plastic and brittle behavior. Moreover, chitin was less sensitive to magnesium stearate and possessed better acetaminophen loading capacity than pregelatinized starch, calcium diphosphate and lactose monohydrate. Chitin showed potential for use as a direct compression excipient. PMID:24528710

Rojas, John; Ciro, Yhors; Correa, Luisa

2014-03-15

304

[Isolation of D-glucosamine from chitin-glucan complexes].  

PubMed

The peculiarities of the acidic hydrolysis of chitin-glucan complexes (CGCs) of higher fungi were studied, and the technology for the isolation and purification of D-(+)-glucosamine hydrochloride of high purity from hydrolysate was developed. The composition, properties, and purity of the product were analyzed by a combination of physicochemical methods. The yield of the final product was 20-60%, depending on the chitin content in CGC samples. The amino sugar obtained was a white crystalline odorless powder readily soluble in water, slightly soluble in 95% ethanol, and insoluble in chloroform and other organic solvents. It corresponds to the standard D-(+)-glucosamine hydrochloride in the main qualitative indicators. PMID:24455869

Artamonova, S D; Sharnina, F F

2013-01-01

305

Determination of chitin in fungi and mycorrhizal roots by an improved HPLC analysis of glucosamine  

Microsoft Academic Search

A method to measure chitin content in fungi and ectomycorrhizal roots with high-performance liquid chromatography (HPLC) was developed. Measurements of fluorescence of 9-fluorenylmethylchloroformate (FMOC-CI) derivatives of glucosamine were made on acid hydrolysates of pure chitin, chitin-root mixtures and fungal-root mixtures. The method was applied on 5 isolates of ectomycorrhizal fungi, and ectomycorrhizal and non-mycorrhizal Pinus sylvestris roots. Interference from amino

Alf Ekblad; Torgny Näsholm

1996-01-01

306

Facile preparation of silver nanoparticles immobilized on chitin nanofiber surfaces to endow antifungal activities.  

PubMed

Silver nanoparticles were prepared on chitin nanofiber surfaces by UV light reduction of silver ions. The chitin nanofibers could be efficient substrates to immobilize silver nanoparticles with stable dispersion states. The dispersion and the nanocomposite film with acrylic resin showed characteristic absorption property in the visible light region due to the effect of the silver nanoparticles. Silver nanoparticles endowed strong antifungal activity to chitin nanofibers. PMID:25498704

Ifuku, Shinsuke; Tsukiyama, Yui; Yukawa, Taisuke; Egusa, Mayumi; Kaminaka, Hironori; Izawa, Hironori; Morimoto, Minoru; Saimoto, Hiroyuki

2015-03-01

307

Chitin utilisation by broilers and its effect on body composition and blood metabolites  

Microsoft Academic Search

1.?Little is known about the ability of farmed poultry to digest chitin and derive nutrients from the ingestion of insects.2.?Commercial chitin derived from crustacean shell waste was found to contain 373?g crude protein, 265?g ash, 23·5?g ether extract, 130?g calcium and 16·4?g phosphorus per kg, on an air-dry basis.3.?It was included in diets at 0, 25, 50 and 75?g chitin

S. M. Hossain; R. Blair

2007-01-01

308

An infrared investigation in relation with chitin and chitosan characterization  

Microsoft Academic Search

The use of infrared spectroscopy for characterization of the composition of chitin and chitosan covering the entire range of degree of acetylation (DA) and a wide variety of raw materials is examined further. The ratio of absorbance bands selected was calibrated using 1H liquid and 13C CP-MAS solid-state NMR as absolute techniques. IR spectra of the structural units of these

J Brugnerotto; J Lizardi; F. M Goycoolea; W Argüelles-Monal; J Desbrières; M Rinaudo

2001-01-01

309

The role of chitin in uranium adsorption by R. arrhizus  

Microsoft Academic Search

In order to further refine and support the uranium biosorption mechanism hypothesis proposed for Rhizopus arrhizus, uranium competitive equilibrium uptake isotherms by chitin were determined at two different solution pH levels and in the presence of different concentrations of competing ions, namely, Cu\\/sup 2 +\\/, Zn\\/sup 2 +\\/, and Fe\\/sup 2 +\\/. The co-ion effect became more pronounced as the

Marios Tsezos

1983-01-01

310

Facile nanofibrillation of chitin derivatives by gas bubbling and ultrasonic treatments in water.  

PubMed

In this paper, we report that nanofiber network structures were constructed from chitin derivatives by gas bubbling and ultrasonic treatments in water. When chitin was first subjected to N2 gas bubbling with ultrasonication in water, the SEM images of the product showed nanofiber network morphology. However, nanofiber network was not re-constructed by the same N2 gas bubbling and ultrasonic treatments after agglomeration. We then have paid attention to an amidine group to provide the agglomeration-nanofibrillation behavior of chitin derivatives. An amidinated chitin was synthesized by the reaction of the amino groups in a partially deacetylated chitin with N,N-dimethylacetamide dimethyl acetal, which was subjected to CO2 gas bubbling and ultrasonic treatments in water to convert into an amidinium chitin by protonation. The SEM images of the product clearly showed nanofiber network morphology. We further examined re-nanofibrillation of the agglomerated material, which was obtained by mixing the nanofibrillated amidinium chitin with water, followed by drying under reduced pressure. Consequently, the material was re-nanofibrillated by N2 gas bubbling with ultrasonication in water owing to electrostatic repulsion between the amidinium groups. Furthermore, deprotonation of the amidinium chitin and re-protonation of the resulting amidinated chitin were conducted by alkaline treatment and CO2 gas bubbling-ultrasonic treatments, respectively. The material showed the agglomeration-nanofibrillation behavior during the processes. PMID:25238127

Tanaka, Kohei; Yamamoto, Kazuya; Kadokawa, Jun-ichi

2014-10-29

311

Nanostructured biocomposite films of high toughness based on native chitin nanofibers and chitosan.  

PubMed

Chitosan is widely used in films for packaging applications. Chitosan reinforcement by stiff particles or fibers is usually obtained at the expense of lowered ductility and toughness. Here, chitosan film reinforcement by a new type of native chitin nanofibers is reported. Films are prepared by casting from colloidal suspensions of chitin in dissolved chitosan. The nanocomposite films are chitin nanofiber networks in chitosan matrix. Characterization is carried out by dynamic light scattering, quartz crystal microbalance, field emission scanning electron microscopy, tensile tests and dynamic mechanical analysis. The polymer matrix nanocomposites were produced in volume fractions of 8, 22, and 56% chitin nanofibers. Favorable chitin-chitosan synergy for colloidal dispersion is demonstrated. Also, lowered moisture sorption is observed for the composites, probably due to the favorable chitin-chitosan interface. The highest toughness (area under stress-strain curve) was observed at 8 vol% chitin content. The toughening mechanisms and the need for well-dispersed chitin nanofibers is discussed. Finally, desired structural characteristics of ductile chitin biocomposites are discussed. PMID:25478558

Mushi, Ngesa E; Utsel, Simon; Berglund, Lars A

2014-01-01

312

Three-dimensional chitin-based scaffolds from Verongida sponges (Demospongiae: Porifera). Part I. Isolation and identification of chitin.  

PubMed

Marine invertebrate organisms including sponges (Porifera) not only provide an abundant source of biologically active secondary metabolites but also inspire investigations to develop biomimetic composites, scaffolds and templates for practical use in materials science, biomedicine and tissue engineering. Here, we presented a detailed study of the structural and physico-chemical properties of three-dimensional skeletal scaffolds of the marine sponges Aiolochroia crassa, Aplysina aerophoba, A. cauliformis, A. cavernicola, and A. fulva (Verongida: Demospongiae). We show that these fibrous scaffolds have a multilayered design and are made of chitin. (13)C solid-state NMR spectroscopy, NEXAFS, and IR spectroscopy as well as chitinase digestion and test were applied in order to unequivocally prove the existence of alpha-chitin in all investigated species. PMID:20471418

Ehrlich, H; Ilan, M; Maldonado, M; Muricy, G; Bavestrello, G; Kljajic, Z; Carballo, J L; Schiaparelli, S; Ereskovsky, A; Schupp, P; Born, R; Worch, H; Bazhenov, V V; Kurek, D; Varlamov, V; Vyalikh, D; Kummer, K; Sivkov, V V; Molodtsov, S L; Meissner, H; Richter, G; Steck, E; Richter, W; Hunoldt, S; Kammer, M; Paasch, S; Krasokhin, V; Patzke, G; Brunner, E

2010-08-01

313

Increased expression of inducible nitric oxide synthase (iNOS) in N-nitrosobis(2-oxopropyl)amine-induced hamster pancreatic carcinogenesis and prevention of cancer development by ONO1714, an iNOS inhibitor  

Microsoft Academic Search

Elevated protein expression of inducible nitric oxide synthase (iNOS) has been observed in human pancreatic cancers and there- fore, iNOS may play important roles in pancreatic carcinogenesis. This was examined in the present study, using an experimental model with N-nitrosobis(2-oxopropyl)amine (BOP)-treated ham- sters. Reverse transcription-polymerase chain reaction analysis demonstrated iNOS expression in a hamster pancreatic cancer cell line as well

Mami Takahashi; Tsukasa Kitahashi; Rikako Ishigamori; Michihiro Mutoh; Masami Komiya; Hidetaka Sato; Yoshihisa Kamanaka; Masao Naka; Takayuki Maruyama; Takashi Sugimura; Keiji Wakabayashi

314

A chalcone synthase/stilbene synthase DNA probe for conifers.  

PubMed

A probe for chalcone synthase (CHS) was generated by PCR using chalcone synthase conserved sequences. The cloned PCR product has high similarity to both chalcone synthase and stilbene synthase sequences. The probe was used to examine the organization of chalcone synthase and stilbene synthase genes in Abies procera, Pinus lambertiana, P. monticola, Picea glauca, P. sitchensis, Pseudostuga menziesii, Taxus brevifolia, and Thuja plicata. A large number of hybridizing bands were found in all species except T. plicata which did not cross hybridize. The hybridization patterns are highly polymorphic between the species and are also polymorphic within several of them. PMID:24166547

Baker, S M; White, E E

1996-05-01

315

ESR studies on reactivity of protein-derived tyrosyl radicals formed by prostaglandin H synthase and ribonucleotide reductase.  

PubMed

Using ESR spectroscopy, the ability of enzyme inhibitors to quench protein-derived tyrosyl radicals was studied in two different enzymes, prostaglandin H synthase and ribonucleotide reductase. The prostaglandin H synthase inhibitors indomethacin, eugenol, and MK-410 effectively prevent the formation of tyrosyl radicals during the oxidation of arachidonic acid by prostaglandin H synthase from ram seminal vesicles. A direct reaction with preformed tyrosyl radicals was observed only with eugenol. The other prostaglandin H synthase inhibitors were ineffective. The ribonucleotide reductase inhibitors hydroxyurea and 4-hydroxyanisole, which effectively inactivate the tyrosyl radical in the active site of ribonucleotide reductase present in tumor cells, exhibit a different reactivity with tyrosyl radicals formed by prostaglandin H synthase. Hydroxyurea quenches preformed tyrosyl radicals in prostaglandin H synthase weakly, whereas 4-hydroxyanisole does not quench tyrosyl radicals in prostaglandin H synthase at all. Eugenol, which quenches preformed prostaglandin H synthase-derived tyrosyl radicals, also quenches the tyrosyl radical in ribonucleotide reductase. The results suggest that the reactivity of protein-linked tyrosyl radicals in ribonucleotide reductase and those formed during prostaglandin H synthase catalysis are very different and have unrelated roles in enzyme catalysis. PMID:8380961

Lassmann, G; Curtis, J; Liermann, B; Mason, R P; Eling, T E

1993-01-01

316

Non-target-site resistance to ALS inhibitors in waterhemp (Amaranthus tuberculatus)  

Technology Transfer Automated Retrieval System (TEKTRAN)

A waterhemp population (MCR) previously characterized as resistant to 4-hyroxyphenylpyruvate dioxygenase (HPPD) and photosystem II (PSII) inhibitors was found to have two different resistance responses to acetolactate synthase (ALS) inhibitors. Plants from the MCR population exhibiting high resistan...

317

Hepoxilin A3 synthase.  

PubMed

Hepoxilins constitute a group of 12S-hydroperoxyeicosatetraenoic acid (12S-HpETE)-derived epoxy-hydroxy fatty acids that have been detected in various cell types and tissues. Although hepoxilin A3 (HXA3) exhibits a myriad of biological activities, its biosynthetic mechanism was not investigated in detail. Here we review the isolation, cloning, and characterization of a leukocyte-type 12S-lipoxygenase (12S-LOX) from rat insulinoma cells RINm5F, which exhibits an intrinsic hepoxilin A3 synthase activity. Confirmation for this observation was achieved by coimmunoprecipitation of HXA3 synthase activity with an anti-leukocyte 12S-LOX antibody, preparation of recombinant rat 12S-LOX enzyme from RINm5F cells, and assay of HXA3 synthase activity therein. Site-directed mutagenesis studies performed on rat 12S-LOX showed that 12-lipoxygenating enzyme species exhibit a strong HXA3 synthase activity that is impaired when the positional specificity of arachidonic acid is altered in favor of 15-lipoxygenation. Inasmuch as cellular glutathione peroxidases (cGPx and PHGPx) and HXA3 synthase compete for the same substrate 12S-HpETE, it can be proposed that the overall activity of glutathione peroxidases, representing the overall peroxide tone, finely tunes the rate of HXA3 formation. PMID:16198304

Nigam, Santosh; Zafiriou, Maria-Patapia

2005-12-01

318

Clinical application of chitin non-woven fabric as wound dressing  

Microsoft Academic Search

A new wound dressing, chitin non-woven fabric was applied in 91 patients for dressing of donor sites, skin graft areas, raw areas under pedicle flaps and those associated with skin abrasion, as well as burns and skin ulcers. Chitin dressing showed excellent results with regard to satisfactory pain relief, adherence to the wound and drying without dissolution or other adverse

Y. Ohshima; K. Nishino; Y. Yonekura; S. Kishimoto; S. Wakabayashi

1987-01-01

319

Soil Bacterial Community Shifts after Chitin Enrichment: An Integrative Metagenomic Approach  

PubMed Central

Chitin is the second most produced biopolymer on Earth after cellulose. Chitin degrading enzymes are promising but untapped sources for developing novel industrial biocatalysts. Hidden amongst uncultivated micro-organisms, new bacterial enzymes can be discovered and exploited by metagenomic approaches through extensive cloning and screening. Enrichment is also a well-known strategy, as it allows selection of organisms adapted to feed on a specific compound. In this study, we investigated how the soil bacterial community responded to chitin enrichment in a microcosm experiment. An integrative metagenomic approach coupling phylochips and high throughput shotgun pyrosequencing was established in order to assess the taxonomical and functional changes in the soil bacterial community. Results indicate that chitin enrichment leads to an increase of Actinobacteria, ?-proteobacteria and ?-proteobacteria suggesting specific selection of chitin degrading bacteria belonging to these classes. Part of enriched bacterial genera were not yet reported to be involved in chitin degradation, like the members from the Micrococcineae sub-order (Actinobacteria). An increase of the observed bacterial diversity was noticed, with detection of specific genera only in chitin treated conditions. The relative proportion of metagenomic sequences related to chitin degradation was significantly increased, even if it represents only a tiny fraction of the sequence diversity found in a soil metagenome. PMID:24278158

Jacquiod, Samuel; Franqueville, Laure; Cécillon, Sébastien; M. Vogel, Timothy; Simonet, Pascal

2013-01-01

320

Hydrogen and oxygen in brine shrimp chitin reflect environmental water and dietary isotopic composition  

Microsoft Academic Search

Hydrogen and oxygen isotope ratios of the common structural biopolymer chitin are a potential recorder of ecological and environmental information, but our understanding of the mechanisms of incorporation of H and O from environmental substrates into chitin is limited. We report the results of a set of experiments in which the isotopic compositions of environmental water and diet were varied

Kristine E. Nielson; Gabriel J. Bowen

2010-01-01

321

Formation of ceramophilic chitin and biohybrid materials enabled by a genetically engineered bifunctional protein.  

PubMed

A bifunctional protein composed of a highly negatively charged oyster shell protein and a chitin-binding domain enabled the formation of biohybrid materials through non-covalent surface modification of chitin nanofibres. The results demonstrate that specific biomolecular interactions offer a route for the formation of biosynthetic materials. PMID:24871427

Malho, Jani-Markus; Heinonen, Hanna; Kontro, Inkeri; Mushi, Ngesa E; Serimaa, Ritva; Hentze, Hans-Peter; Linder, Markus B; Szilvay, Géza R

2014-07-14

322

Cadmium removal from aqueous solutions by chitin: kinetic and equilibrium studies  

Microsoft Academic Search

A fundamental investigation on the removal of cadmium ions from aqueous solutions by chitin was conducted in batch conditions. Kinetic data and equilibrium removal isotherms were measured. The influence of different experimental parameters such as time contact, initial concentration of cadmium, chitin mass, particles size, agitation speed, temperature and the nature of cadmium salt, on the kinetics of cadmium removal

B Benguella; H Benaissa

2002-01-01

323

Application of chitin- and chitosan-based materials for enzyme immobilizations: a review  

Microsoft Academic Search

As functional materials, chitin and chitosan offer a unique set of characteristics: biocompatibility, biodegradability to harmless products, nontoxicity, physiological inertness, antibacterial properties, heavy metal ions chelation, gel forming properties and hydrophilicity, and remarkable affinity to proteins. Owing to these characteristics, chitin- and chitosan-based materials, as yet underutilized, are predicted to be widely exploited in the near future especially in environmentally

Barbara Krajewska

2004-01-01

324

Interplay of liquid-crystallinity and gelation: the two limiting cases of chitin and actin suspensions  

E-print Network

Liquid level Isotropic phase I/LC interface Liquid-crystalline phase Textures in polarised lightInterplay of liquid-crystallinity and gelation: the two limiting cases of chitin and actin-elastic suspensions of rod-like polymers always really liquid-crystalline ? #12;Chitin (commercial product) Acid

Paris-Sud 11, Université de

325

Chitin-based renewable materials from marine sponges for uranium adsorption.  

PubMed

Marine sponges of the order Verongida form three-dimensional networks of fibrous chitin, which can easily be extracted. In the hydrated state, these networks are flexible, mechanically stable and can be cut or pressed into any desired form. Here, we show for the first time that chitin-based networks of sponge origin are useful for effective uranium adsorption. They adsorb uranium from solution with a higher adsorption capacity than many other chitinous sorbents. Up to 288 mg/g could be achieved. Solid-state NMR, infrared, and Raman spectroscopy indicated that the uranyl is bound to the chitin by weak interactions. 90% of the uranyl could be desorbed using diluted hydrochloric acid. Uranium adsorption and desorption did not result in any destruction of the chitin-based material. PMID:23218358

Schleuter, Dorothea; Günther, Alix; Paasch, Silvia; Ehrlich, Hermann; Kljaji?, Zoran; Hanke, Thomas; Bernhard, Gert; Brunner, Eike

2013-01-30

326

Control of Soybean Cyst Nematode by Chitinolytic Bacteria with Chitin Substrate  

PubMed Central

Sixty-four chitinolytic bacterial isolates from soybean fields in Arkansas were tested for suppression of soybean cyst nematode (Heterodera glycines) in a heat-treated silt loam soil amended with 0.6% (w/w) chitin in a greenhouse. Five isolates consistently reduced numbers of H. glycines compared to controls receiving neither chitin nor bacteria, or only chitin. Four of the five isolates interacted with the chitin substrate to enhance its effectiveness in reducing numbers of the nematode. The size of the clear-zone produced by some of the isolates in colloidal chitin medium, an indication of chitinolytic activity in vitro, was not related to suppression of nematode numbers in soil. PMID:19270991

Tian, Honglin; Riggs, Robert D.; Crippen, Devany L.

2000-01-01

327

Effects of supercritical water and mechanochemical grinding treatments on physicochemical properties of chitin.  

PubMed

This study examined the effects of a combined pretreatment with supercritical water and mechanochemical grinding with a ball mill on the physicochemical properties of chitin and its enzymatic degradation. Following pretreatment with a combination of supercritical water and grinding, chitin had a lower mean molecular weight, a lower crystallinity index, a lower crystallite size, greater d-spacing, weaker hydrogen bonds, and the amide group was more exposed compared with untreated chitin. These properties increased the hydrophilicity of the chitin and enhanced its enzymatic degradation. The N,N'-diacetylchitobiose (GlcNAc)(2) yield after enzymatic degradation of chitin following pretreatment with supercritical water (400 °C, 1 min) and grinding (800 rpm, 10 min) was 93%, compared with 5% without any treatment, 37% with supercritical water pretreatment alone (400 °C, 1 min), and 60% with grinding alone (800 rpm, 30 min). PMID:23399191

Osada, Mitsumasa; Miura, Chika; Nakagawa, Yuko S; Kaihara, Mikio; Nikaido, Mitsuru; Totani, Kazuhide

2013-02-15

328

Blends of chitin and chitosan with polyamide 66  

SciTech Connect

For several years, intense interest has been focused on polymer blends in which both components are synthetic polymers. However, few studies have been made on blends in which one component is chitin (QA), or chitosan (QN), the most abundant natural polymers after cellulose. Its chemical structure, based in partially acetilated {beta}-aminosaccharide units, permits the formation of natural blends with proteins and inorganic salts were the intermolecular hydrogen bonds play an important role. The choice of a partner for these natural polymers was made expecting strong interaction between the two polymers. For this reason, on this work, polyamide 66 (P66), has been chosen.

Gonzalez, V. [CIQA, Saltillo (Mexico); Guerrero, C. [DIMAT-FIME-UANL (Mexico)

1996-12-31

329

Microbial responses to chitin and chitosan in oxic and anoxic agricultural soil slurries  

NASA Astrophysics Data System (ADS)

Microbial degradation of chitin in soil substantially contributes to carbon cycling in terrestrial ecosystems. Chitin is globally the second most abundant biopolymer after cellulose and can be deacetylated to chitosan or can be hydrolyzed to N,N'-diacetylchitobiose and oligomers of N-acetylglucosamine by aerobic and anaerobic microorganisms. Which pathway of chitin hydrolysis is preferred by soil microbial communities is unknown. Supplementation of chitin stimulated microbial activity under oxic and anoxic conditions in agricultural soil slurries, whereas chitosan had no effect. Thus, the soil microbial community likely was more adapted to chitin as a substrate. In addition, this finding suggested that direct hydrolysis of chitin was preferred to the pathway that starts with deacetylation. Chitin was apparently degraded by aerobic respiration, ammonification, and nitrification to carbon dioxide and nitrate under oxic conditions. When oxygen was absent, fermentation products (acetate, butyrate, propionate, hydrogen, and carbon dioxide) and ammonia were detected, suggesting that butyric and propionic acid fermentation, along with ammonification, were likely responsible for anaerobic chitin degradation. In total, 42 different chiA genotypes were detected of which twenty were novel at an amino acid sequence dissimilarity of less than 50%. Various chiA genotypes responded to chitin supplementation and affiliated with a novel deep-branching bacterial chiA genotype (anoxic conditions), genotypes of Beta- and Gammaproteobacteria (oxic and anoxic conditions), and Planctomycetes (oxic conditions). Thus, this study provides evidence that detected chitinolytic bacteria were catabolically diverse and occupied different ecological niches with regard to oxygen availability enabling chitin degradation under various redox conditions on community level.

Wieczorek, A. S.; Hetz, S. A.; Kolb, S.

2014-06-01

330

Microbial responses to chitin and chitosan in oxic and anoxic agricultural soil slurries  

NASA Astrophysics Data System (ADS)

Chitin is the second most abundant biopolymer in terrestrial ecosystems and is subject to microbial degradation. Chitin can be deacetylated to chitosan or can be hydrolyzed to N,N'-diacetylchitobiose and oligomers of N-acetylglucosamine by aerobic and anaerobic microorganisms. Which pathway of chitin hydrolysis is preferred by soil microbial communities has previously been unknown. Supplementation of chitin stimulated microbial activity under oxic and anoxic conditions in agricultural soil slurries, whereas chitosan had no effect. Thus, the soil microbial community likely was more adapted to chitin as a substrate. In addition, this finding suggested that direct hydrolysis of chitin was preferred to the pathway that starts with deacetylation. Chitin was apparently degraded by aerobic respiration, ammonification, and nitrification to carbon dioxide and nitrate under oxic conditions. When oxygen was absent, fermentation products (acetate, butyrate, propionate, hydrogen, carbon dioxide) and ammonia were detected, suggesting that butyric and propionic acid fermentation were along with ammonification likely responsible for apparent anaerobic chitin degradation. In total, 42 different chiA genotypes were detected of which twenty were novel at an amino acid sequence dissimilarity of >50%. Various chiA genotypes responded to chitin supplementation and affiliated with a novel deep-branching bacterial chiA genotype (anoxic conditions), genotypes of Beta- and Gammaproteobacteria (oxic and anoxic conditions), and Planctomycetes (oxic conditions). Thus, this study provides evidence that detected chitinolytic bacteria were catabolically diverse and occupied different ecological niches with regard to oxygen availability enabling chitin degradation under various redox conditions at the level of the community.

Wieczorek, A. S.; Hetz, S. A.; Kolb, S.

2014-02-01

331

Geranyl diphosphate synthase from mint  

DOEpatents

A cDNA encoding geranyl diphosphate synthase from peppermint has been isolated and sequenced, and the corresponding amino acid sequence has been determined. Accordingly, an isolated DNA sequence (SEQ ID No:1) is provided which codes for the expression of geranyl diphosphate synthase (SEQ ID No:2) from peppermint (Mentha piperita). In other aspects, replicable recombinant cloning vehicles are provided which code for geranyl diphosphate synthase or for a base sequence sufficiently complementary to at least a portion of the geranyl diphosphate synthase DNA or RNA to enable hybridization therewith (e.g., antisense geranyl diphosphate synthase RNA or fragments of complementary geranyl diphosphate synthase DNA which are useful as polymerase chain reaction primers or as probes for geranyl diphosphate synthase or related genes). In yet other aspects, modified host cells are provided that have been transformed, transfected, infected and/or injected with a recombinant cloning vehicle and/or DNA sequence encoding geranyl diphosphate synthase. Thus, systems and methods are provided for the recombinant expression of geranyl diphosphate synthase that may be used to facilitate the production, isolation and purification of significant quantities of recombinant geranyl diphosphate synthase for subsequent use, to obtain expression or enhanced expression of geranyl diphosphate synthase in plants in order to enhance the production of monoterpenoids, to produce geranyl diphosphate in cancerous cells as a precursor to monoterpenoids having anti-cancer properties or may be otherwise employed for the regulation or expression of geranyl diphosphate synthase or the production of geranyl diphosphate. 5 figs.

Croteau, R.B.; Wildung, M.R.; Burke, C.C.; Gershenzon, J.

1999-03-02

332

Geranyl diphosphate synthase from mint  

DOEpatents

A cDNA encoding geranyl diphosphate synthase from peppermint has been isolated and sequenced, and the corresponding amino acid sequence has been determined. Accordingly, an isolated DNA sequence (SEQ ID No:1) is provided which codes for the expression of geranyl diphosphate synthase (SEQ ID No:2) from peppermint (Mentha piperita). In other aspects, replicable recombinant cloning vehicles are provided which code for geranyl diphosphate synthase or for a base sequence sufficiently complementary to at least a portion of the geranyl diphosphate synthase DNA or RNA to enable hybridization therewith (e.g., antisense geranyl diphosphate synthase RNA or fragments of complementary geranyl diphosphate synthase DNA which are useful as polymerase chain reaction primers or as probes for geranyl diphosphate synthase or related genes). In yet other aspects, modified host cells are provided that have been transformed, transfected, infected and/or injected with a recombinant cloning vehicle and/or DNA sequence encoding geranyl diphosphate synthase. Thus, systems and methods are provided for the recombinant expression of geranyl diphosphate synthase that may be used to facilitate the production, isolation and purification of significant quantities of recombinant geranyl diphosphate synthase for subsequent use, to obtain expression or enhanced expression of geranyl diphosphate synthase in plants in order to enhance the production of monoterpenoids, to produce geranyl diphosphate in cancerous cells as a precursor to monoterpenoids having anti-cancer properties or may be otherwise employed for the regulation or expression of geranyl diphosphate synthase or the production of geranyl diphosphate.

Croteau, Rodney Bruce (Pullman, WA); Wildung, Mark Raymond (Colfax, WA); Burke, Charles Cullen (Moscow, ID); Gershenzon, Jonathan (Jena, DE)

1999-01-01

333

Inhibition of glucosylceramide synthase stimulates autophagy flux in neurons.  

PubMed

Aggregate-prone mutant proteins, such as ?-synuclein and huntingtin, play a prominent role in the pathogenesis of various neurodegenerative disorders; thus, it has been hypothesized that reducing the aggregate-prone proteins may be a beneficial therapeutic strategy for these neurodegenerative disorders. Here, we identified two previously described glucosylceramide (GlcCer) synthase inhibitors, DL-threo-1-Phenyl-2-palmitoylamino-3-morpholino-1-propanol and Genz-123346(Genz), as enhancers of autophagy flux. We also demonstrate that GlcCer synthase inhibitors exert their effects on autophagy by inhibiting AKT-mammalian target of rapamycin (mTOR) signaling. More importantly, siRNA knock down of GlcCer synthase had the similar effect as pharmacological inhibition, confirming the on-target effect. In addition, we discovered that inhibition of GlcCer synthase increased the number and size of lysosomal/late endosomal structures. Although inhibition of GlcCer synthase decreases levels of mutant ?-synuclein in neurons, it does so, according to our data, through autophagy-independent mechanisms. Our findings demonstrate a direct link between glycosphingolipid biosynthesis and autophagy in primary neurons, which may represent a novel pathway with potential therapeutic value for the treatment of Parkinson's disease. Inhibition of GlcCer synthase enhances autophagy by inhibiting AKT-mTOR signaling, and increases the number and size of lysosomal/late endosomal structures. Furthermore, inhibition of GlcCer synthase decreased levels of mutant ?-synuclein in neurons, which may represent a potential therapeutic target for Parkinson's disease. PMID:24494600

Shen, Wei; Henry, Anastasia G; Paumier, Katrina L; Li, Li; Mou, Kewa; Dunlop, John; Berger, Zdenek; Hirst, Warren D

2014-06-01

334

Tapentadol and nitric oxide synthase systems.  

PubMed

Tapentadol, a new analgesic drug with a dual mechanism of action (?-opioid receptor agonism and norepinephrine reuptake inhibition), is indicated for the treatment of moderate to severe acute and chronic pain. In this paper, the possible additional involvement of the nitric oxide synthase (NOS) system in the antinociceptive activity of tapentadol was investigated using an unspecific inhibitor of NOS, L-NOArg, a relatively specific inhibitor of neuronal NOS, 7-NI, a relatively selective inhibitor of inducible NOS, L-NIL, and a potent inhibitor of endothelial NOS, L-NIO. Tapentadol (1-10?mg/kg, intraperitoneal) increased the threshold for mechanical (Randall-Selitto test) and thermal (tail-flick test) nociceptive stimuli in a dose-dependent manner. All four NOS inhibitors, administered intraperitoneally in the dose range 0.1-10?mg/kg, potentiated the analgesic action of tapentadol at a low dose of 2?mg/kg in both models of pain. We conclude that NOS systems participate in tapentadol analgesia. PMID:25485639

Bujalska-Zadro?ny, Magdalena; Woli?ska, Renata; G?si?ska, Emilia; Nagraba, Lukasz

2014-12-01

335

Fungal Chitin Dampens Inflammation through IL-10 Induction Mediated by NOD2 and TLR9 Activation  

PubMed Central

Chitin is an essential structural polysaccharide of fungal pathogens and parasites, but its role in human immune responses remains largely unknown. It is the second most abundant polysaccharide in nature after cellulose and its derivatives today are widely used for medical and industrial purposes. We analysed the immunological properties of purified chitin particles derived from the opportunistic human fungal pathogen Candida albicans, which led to the selective secretion of the anti-inflammatory cytokine IL-10. We identified NOD2, TLR9 and the mannose receptor as essential fungal chitin-recognition receptors for the induction of this response. Chitin reduced LPS-induced inflammation in vivo and may therefore contribute to the resolution of the immune response once the pathogen has been defeated. Fungal chitin also induced eosinophilia in vivo, underpinning its ability to induce asthma. Polymorphisms in the identified chitin receptors, NOD2 and TLR9, predispose individuals to inflammatory conditions and dysregulated expression of chitinases and chitinase-like binding proteins, whose activity is essential to generate IL-10-inducing fungal chitin particles in vitro, have also been linked to inflammatory conditions and asthma. Chitin recognition is therefore critical for immune homeostasis and is likely to have a significant role in infectious and allergic disease. Authors Summary Chitin is the second most abundant polysaccharide in nature after cellulose and an essential component of the cell wall of all fungal pathogens. The discovery of human chitinases and chitinase-like binding proteins indicates that fungal chitin is recognised by cells of the human immune system, shaping the immune response towards the invading pathogen. We show that three immune cell receptors– the mannose receptor, NOD2 and TLR9 recognise chitin and act together to mediate an anti-inflammatory response via secretion of the cytokine IL-10. This mechanism may prevent inflammation-based damage during fungal infection and restore immune balance after an infection has been cleared. By increasing the chitin content in the cell wall pathogenic fungi may influence the immune system in their favour, by down-regulating protective inflammatory immune responses. Furthermore, gene mutations and dysregulated enzyme activity in the described chitin recognition pathway are implicated in inflammatory conditions such as Crohn's Disease and asthma, highlighting the importance of the discovered mechanism in human health. PMID:24722226

Wagener, Jeanette; Malireddi, R. K. Subbarao; Lenardon, Megan D.; Köberle, Martin; Vautier, Simon; MacCallum, Donna M.; Biedermann, Tilo; Schaller, Martin; Netea, Mihai G.; Kanneganti, Thirumala-Devi; Brown, Gordon D.; Brown, Alistair J. P.; Gow, Neil A. R.

2014-01-01

336

Inhibition of ATP Synthase by Chlorinated Adenosine Analogue  

PubMed Central

8-Chloroadenosine (8-Cl-Ado) is a ribonucleoside analogue that is currently in clinical trial for chronic lymphocytic leukemia. Based on the decline in cellular ATP pool following 8-Cl-Ado treatment, we hypothesized that 8-Cl-ADP and 8-Cl-ATP may interfere with ATP synthase, a key enzyme in ATP production. Mitochondrial ATP synthase is composed of two major parts; FO intermembrane base and F1 domain, containing ? and ? subunits. Crystal structures of both ? and ? subunits that bind to the substrate, ADP, are known in tight binding (?dp?dp) and loose binding (?tp?tp) states. Molecular docking demonstrated that 8-Cl-ADP/8-Cl-ATP occupied similar binding modes as ADP/ATP in the tight and loose binding sites of ATP synthase, respectively, suggesting that the chlorinated nucleotide metabolites may be functional substrates and inhibitors of the enzyme. The computational predictions were consistent with our whole cell biochemical results. Oligomycin, an established pharmacological inhibitor of ATP synthase, decreased both ATP and 8-Cl-ATP formation from exogenous substrates, however, did not affect pyrimidine nucleoside analogue triphosphate accumulation. Synthesis of ATP from ADP was inhibited in cells loaded with 8-Cl-ATP. These biochemical studies are in consent with the computational modeling; in the ?tp?tp state 8-Cl-ATP occupies similar binding as ANP, a non-hydrolyzable ATP mimic that is a known inhibitor. Similarly, in the substrate binding site (?dp?dp) 8-Cl-ATP occupies a similar position as ATP mimic ADP-BeF3 ?. Collectively, our current work suggests that 8-Cl-ADP may serve as a substrate and the 8-Cl-ATP may be an inhibitor of ATP synthase. PMID:19477165

Chen, Lisa S.; Nowak, Billie J.; Ayres, Mary L.; Krett, Nancy L.; Rosen, Steven T.; Zhang, Shuxing; Gandhi, Varsha

2009-01-01

337

Use of Chitin for Controlling Heterodera avenae and Tylenchulus semipenetrans.  

PubMed

The nematicidal effect of chitin, relative to other pesticides, was evaluated against two plant-parasitic nematodes, Heterodera avenae and Tylenchulus semipenetrans. Wheat seedlings, grown in soils artificially or naturally infested with H. avenae, were treated with 0.4% (w/w) ClandoSan (CLA) prepared from crustacean chitin, aldicarb (Temik 15G), or ethylene dibromide (EDB 90EC). The CLA treatment significantly increased wheat straw, ear, and average grain dry weights of nematode-infected plants, compared with the other two treatments. In an experiment covering two consecutive seasons, all three treatments reduced the number of cysts in the soil by 60%. In a one-season experiment, CLA reduced the number of cysts by 51% and aldicarb or EDB reduced cyst number by about 40%. A reduction of 50-90% in T. semipenetrans population densities on roots of two citrus rootstocks was recorded following an application of 0.2% (w/w) CLA to the soil. PMID:19287630

Spiegel, Y; Cohn, E; Chet, I

1989-07-01

338

Use of Chitin for Controlling Heterodera avenae and Tylenchulus semipenetrans  

PubMed Central

The nematicidal effect of chitin, relative to other pesticides, was evaluated against two plant-parasitic nematodes, Heterodera avenae and Tylenchulus semipenetrans. Wheat seedlings, grown in soils artificially or naturally infested with H. avenae, were treated with 0.4% (w/w) ClandoSan (CLA) prepared from crustacean chitin, aldicarb (Temik 15G), or ethylene dibromide (EDB 90EC). The CLA treatment significantly increased wheat straw, ear, and average grain dry weights of nematode-infected plants, compared with the other two treatments. In an experiment covering two consecutive seasons, all three treatments reduced the number of cysts in the soil by 60%. In a one-season experiment, CLA reduced the number of cysts by 51% and aldicarb or EDB reduced cyst number by about 40%. A reduction of 50-90% in T. semipenetrans population densities on roots of two citrus rootstocks was recorded following an application of 0.2% (w/w) CLA to the soil. PMID:19287630

Spiegel, Y.; Cohn, E.; Chet, I.

1989-01-01

339

Pyrolysis GC/MS and IR spectroscopy in chitin analysis of molluscan shells.  

PubMed

Chitin is an insoluble component in the shells of several molluscan species. It is thought to play important roles, in biomineralization and shell structure. To date, however, reports are scarce and sometimes contradictory, and suffer from methodological problems. Only in a single cephalopod species has the chitin been identified as beta-chitin. We present data on chitin occurrence in 22 species of shell-bearing Mollusca (Conchifera) and Polyplacophora, including the first evidence for scaphopods, based on pyrolysis gas chromatography, mass spectrometry (GC-MS), and infrared spectroscopy (IR). Pyrolysis GC-MS detected chitin in every tested member of the Conchifera. IR spectroscopy before and after chitinase treatment revealed at least three distinct patterns of peak changes. The contents of the insoluble shell organics included not only chitin and proteins, but also insoluble polysaccharides, e.g., glucan. We conclude that chitin was present in the last common ancestor of the Conchifera and that its abundance in the shell matrix depends on the differentiation of the shell. PMID:19129649

Furuhashi, Takeshi; Beran, Anton; Blazso, Marianne; Czegeny, Zsuzsanna; Schwarzinger, Clemens; Steiner, Gerhard

2009-01-01

340

Green Conversion of Agroindustrial Wastes into Chitin and Chitosan by Rhizopus arrhizus and Cunninghamella elegans Strains  

PubMed Central

This article sets out a method for producing chitin and chitosan by Cunninghamella elegans and Rhizopus arrhizus strains using a green metabolic conversion of agroindustrial wastes (corn steep liquor and molasses). The physicochemical characteristics of the biopolymers and antimicrobial activity are described. Chitin and chitosan were extracted by alkali-acid treatment, and characterized by infrared spectroscopy, viscosity and X-ray diffraction. The effectiveness of chitosan from C. elegans and R. arrhizus in inhibiting the growth of Listeria monocytogenes, Staphylococcus aureus, Pseudomonas aeruginosa, Salmonella enterica, Escherichia coli and Yersinia enterocolitica were evaluated by determining the minimum inhibitory concentrations (MIC) and the minimum bactericidal concentrations (MBC). The highest production of biomass (24.60 g/L), chitin (83.20 mg/g) and chitosan (49.31 mg/g) was obtained by R. arrhizus. Chitin and chitosan from both fungi showed a similar degree of deacetylation, respectively of 25% and 82%, crystallinity indices of 33.80% and 32.80% for chitin, and 20.30% and 17.80% for chitosan. Both chitin and chitosan presented similar viscosimetry of 3.79–3.40 cP and low molecular weight of 5.08 × 103 and 4.68 × 103 g/mol. They both showed identical MIC and MBC for all bacteria assayed. These results suggest that: agricultural wastes can be produced in an environmentally friendly way; chitin and chitosan can be produced economically; and that chitosan has antimicrobial potential against pathogenic bacteria. PMID:24853288

Berger, Lúcia Raquel Ramos; Stamford, Thayza Christina Montenegro; Stamford-Arnaud, Thatiana Montenegro; de Alcântara, Sergio Roberto Cabral; da Silva, Antonio Cardoso; da Silva, Adamares Marques; do Nascimento, Aline Elesbão; de Campos-Takaki, Galba Maria

2014-01-01

341

Salmonella biofilm formation on Aspergillus niger involves cellulose--chitin interactions.  

PubMed

Salmonella cycles between host and nonhost environments, where it can become an active member of complex microbial communities. The role of fungi in the environmental adaptation of enteric pathogens remains relatively unexplored. We have discovered that S. enterica Typhimurium rapidly attaches to and forms biofilms on the hyphae of the common fungus, Aspergillus niger. Several Salmonella enterica serovars displayed a similar interaction, whereas other bacterial species were unable to bind to the fungus. Bacterial attachment to chitin, a major constituent of fungal cell walls, mirrored this specificity. Pre-incubation of S. Typhimurium with N-acetylglucosamine, the monomeric component of chitin, reduced binding to chitin beads by as much as 727-fold and inhibited attachment to A. niger hyphae considerably. A cellulose-deficient mutant of S. Typhimurium failed to attach to chitin beads and to the fungus. Complementation of this mutant with the cellulose operon restored binding to chitin beads to 79% of that of the parental strain and allowed for attachment and biofilm formation on A. niger, indicating that cellulose is involved in bacterial attachment to the fungus via the chitin component of its cell wall. In contrast to cellulose, S. Typhimurium curli fimbriae were not required for attachment and biofilm development on the hyphae but were critical for its stability. Our results suggest that cellulose-chitin interactions are required for the production of mixed Salmonella-A. niger biofilms, and support the hypothesis that encounters with chitinaceous alternate hosts may contribute to the ecological success of human pathogens. PMID:22003399

Brandl, Maria T; Carter, Michelle Q; Parker, Craig T; Chapman, Matthew R; Huynh, Steven; Zhou, Yaguang

2011-01-01

342

Electron microscopic localization of chitin using colloidal gold labelled with wheat germ agglutinin.  

PubMed

The lectin wheat germ agglutinin (WGA) has a binding site which is able to bind a sequence of three N-acetyl-glucosamine residues. Therefore, it has a very strong affinity for the polymers of this sugar, especially chitin. Colloidal gold can be labelled with WGA and used as a specific electron-dense marker for the electron-microscopic localization of chitin. The specificity of the WGA-gold binding can be checked by competitive inhibition with 5-10 mM triacetyl chitotriose. The reliability of this method was tested in three species. In the formation zone of the radula of the snail, Biomphalaria glabrata Say, chitin or chitin precursors were localized in vesicles of the odontoblasts, outside the extremely long microvilli of odontoblasts and in the newly formed teeth. The inner peritrophic envelope of the earwig, Forficula auricularia L., is characterized by an orthogonal texture of bundles of microfibrils that are thought to contain chitin. The presence of chitin was proved using the present method. In the peritrophic membranes of the blowfly, Calliphora erythrocephala Meigen, it was possible to differentiate between chitin and glycoproteins which have N-acetylglucosamine residues. PMID:3754855

Peters, W; Latka, I

1986-01-01

343

Construction of chitin/PVA composite hydrogels with jellyfish gel-like structure and their biocompatibility.  

PubMed

High strength chitin/poly(vinyl alcohol) (PVA) composite hydrogels (RCP) were constructed by adding PVA into chitin dissolved in a NaOH/urea aqueous solution, and then by cross-linking with epichlorohydrin (ECH) and freezing-thawing process. The RCP hydrogels were characterized by field emission scanning electron microscopy, FTIR, differential scanning calorimetry, solid-state (13)C NMR, wide-angle X-ray diffraction, and compressive test. The results revealed that the repeated freezing/thawing cycles induced the bicrosslinked networks consisted of chitin and PVA crystals in the composite gels. Interestingly, a jellyfish gel-like structure occurred in the RCP75 gel with 25 wt % PVA content in which the amorphous and crystalline PVA were immobilized tightly in the chitin matrix through hydrogen bonding interaction. The freezing/thawing cycles played an important role in the formation of the layered porous PVA networks and the tight combining of PVA with the pore wall of chitin. The mechanical properties of RCP75 were much higher than the other RCP gels, and the compressive strength was 20× higher than that of pure chitin gels, as a result of broadly dispersing stress caused by the orderly multilayered networks. Furthermore, the cell culture tests indicated that the chitin/PVA composite hydrogels exhibited excellent biocompatibility and safety, showing potential applications in the field of tissue engineering. PMID:25077674

He, Meng; Wang, Zhenggang; Cao, Yan; Zhao, Yanteng; Duan, Bo; Chen, Yun; Xu, Min; Zhang, Lina

2014-09-01

344

Characterization of chitin and its hydrolysis to GlcNAc and GlcN.  

PubMed

Proton NMR spectra of chitin dissolved in concentrated and deuterated hydrochloric acid (DCl) were found to be a simple and powerful method for identifying chitin from samples of biological origin. During the first hour after dissolving chitin in concentrated DCl (25 degrees C), insignificant de-N-acetylation occurred, meaning that the fraction of acetylated units (FA) of chitin could be determined. FA of demineralized shrimp shell samples treated with 1 M NaOH at 95 degrees C for 1-24 h were determined and were found to decrease linearly with time from 0.96 to 0.91 during the treatment with NaOH. Extrapolation to zero time suggested that chitin from shrimp shells has a FA of 0.96, that is, contains a small but significant fraction of de-N-acetylated units. Proton NMR spectra of chitin ( FA = 0.96) dissolved in concentrated DCl were obtained as a function of time until the samples were almost quantitatively hydrolyzed to the monomer glucosamine (GlcN). The initial phase of the reaction involves mainly depolymerization of the chitin chains, resulting in that almost 90% (molar fraction) of the chitin is converted to the monomer N-acetyl-glucosamine (GlcNAc).Thus, effective conversion of chitin to GlcNAc in concentrated acid is reported for the first time. GlcNAc is then further de-N-acetylated to GlcN. A new theoretical model was developed to simulate the experimental data of the kinetics of hydrolysis of chitin in concentrated acid. The model uses three different rate constants; two for the hydrolysis of the glycosidic linkages following an N-acetylated or a de-N-acetylated sugar unit and one for the de-N-acetylation reaction. The three rate constants were estimated by fitting model data to experimental results. The rate of hydrolysis of a glycosidic linkage following an N-acetylated unit was found to be 54 times higher as compared to the rate of de-N-acetylation and 115 times higher than the rate of hydrolysis of a glycosidic linkage following a de-N-acetylated unit. Two chitin samples with different F A values (0.96 and 0.70) were incubated in concentrated DCl until the samples were converted to the maximum yield of GlcNAc and the oligomer composition analyzed, showing that the maximum yield of GlcNAc was much higher when prepared from the chitin with the highest F A value. PMID:18540645

Einbu, Aslak; Vårum, Kjell M

2008-07-01

345

Chitin-based scaffolds are an integral part of the skeleton of the marine demosponge Ianthella basta.  

PubMed

The skeletons of demosponges, such as Ianthella basta, are known to be a composite material containing organic constituents. Here, we show that a filigree chitin-based scaffold is an integral component of the I. basta skeleton. These chitin-based scaffolds can be isolated from the sponge skeletons using an isolation and purification technique based on treatment with alkaline solutions. Solid-state (13)C NMR, Raman, and FT-IR spectroscopies, as well as chitinase digestion, reveal that the isolated material indeed consists of chitin. The morphology of the scaffolds has been determined by light and electron microscopy. It consists of cross-linked chitin fibers approximately 40-100 nm in diameter forming a micro-structured network. The overall shape of this network closely resembles the shape of the integer sponge skeleton. Solid-state (13)C NMR spectroscopy was used to characterize the sponge skeleton on a molecular level. The (13)C NMR signals of the chitin-based scaffolds are relatively broad, indicating a high amount of disordered chitin, possibly in the form of surface-exposed molecules. X-ray diffraction confirms that the scaffolds isolated from I. basta consist of partially disordered and loosely packed chitin with large surfaces. The spectroscopic signature of these chitin-based scaffolds is closer to that of alpha-chitin than beta-chitin. PMID:19567270

Brunner, E; Ehrlich, H; Schupp, P; Hedrich, R; Hunoldt, S; Kammer, M; Machill, S; Paasch, S; Bazhenov, V V; Kurek, D V; Arnold, T; Brockmann, S; Ruhnow, M; Born, R

2009-12-01

346

Chitin-based scaffolds are an integral part of the skeleton of the marine demosponge Ianthella basta  

PubMed Central

The skeletons of demosponges, such as Ianthella basta, are known to be a composite material containing organic constituents. Here, we show that a filigree chitin-based scaffold is an integral component of the I. basta skeleton. These chitin-based scaffolds can be isolated from the sponge skeletons using an isolation and purification technique based on treatment with alkaline solutions. Solid-state 13C NMR, Raman, and FT-IR spectroscopies, as well as chitinase digestion, reveal that the isolated material indeed consists of chitin. The morphology of the scaffolds has been determined by light and electron microscopy. It consists of cross-linked chitin fibers approximately 40–100 nm in diameter forming a micro-structured network. The overall shape of this network closely resembles the shape of the integer sponge skeleton. Solid-state 13C NMR spectroscopy was used to characterize the sponge skeleton on a molecular level. The 13C NMR signals of the chitin-based scaffolds are relatively broad, indicating a high amount of disordered chitin, possibly in the form of surface-exposed molecules. X-ray diffraction confirms that the scaffolds isolated from I. basta consist of partially disordered and loosely packed chitin with large surfaces. The spectroscopic signature of these chitin-based scaffolds is closer to that of ?-chitin than ?-chitin. PMID:19567270

Brunner, E.; Ehrlich, H.; Schupp, P.; Hedrich, R.; Hunoldt, S.; Kammer, M.; Machill, S.; Paasch, S.; Bazhenov, V.V.; Kurek, D.V.; Arnold, T.; Brockmann, S.; Ruhnow, M.; Born, R.

2010-01-01

347

Single Agents with Designed Combination Chemotherapy Potential: Synthesis and Evaluation of Substituted Pyrimido[4,5-b]indoles as Receptor Tyrosine Kinase and Thymidylate Synthase Inhibitors and as Antitumor Agents  

PubMed Central

Combinations of antiangiogenic agents (AAs) with cytotoxic agents have shown significant promise and several such clinical trials are currently underway. We have designed, synthesized and evaluated two compounds that each inhibit vascular endothelial growth factor receptor-2 (VEGFR-2) and platelet derived growth factor receptor-beta (PDGFR-?) for antiangiogenic effects and also inhibit human thymidylate synthase (hTS) for cytotoxic effects in single agents. The synthesis of these compounds involved the nucleophilic displacement of the common intermediate 5-chloro-9H-pyrimido[4,5-b]indole-2,4-diamine with appropriate benzenethiols. The inhibitory potency of both these single agents against VEGFR-2, PDGFR-? and hTS is better than or close to standards. In a COLO-205 xenograft mouse model one of the analogs significantly decreased tumor growth (TGI = 76% at 35 mg/kg), liver metastases and tumor blood vessels compared to a standard drug and to control and thus demonstrated potent tumor growth inhibition, inhibition of metastasis and antiangiogenic effects in vivo. These compounds afford combination chemotherapeutic potential in single agents. PMID:20092323

Gangjee, Aleem; Zaware, Nilesh; Raghavan, Sudhir; Ihnat, Michael; Shenoy, Satyendra; Kisliuk, Roy L.

2010-01-01

348

MICROBIAL FERMENTATION OF ABUNDANT BIOPOLYMERS: CELLULOSE AND CHITIN  

SciTech Connect

Our research has dealt with seven major areas of investigation: i) characterization of cellulolytic members of microbial consortia, with special attention recently given to Clostridium phytofermentans, a bacterium that decomposes cellulose and produces uncommonly large amounts of ethanol, ii) investigations of the chitinase system of Cellulomonas uda; including the purification and characterization of ChiA, the major component of this enzyme system, iii) molecular cloning, sequence and structural analysis of the gene that encodes ChiA in C. uda, iv) biofilm formation by C. uda on nutritive surfaces, v) investigations of the effects of humic substances on cellulose degradation by anaerobic cellulolytic microbes, vi) studies of nitrogen metabolism in cellulolytic anaerobes, and vii) understanding the molecular architecture of the multicomplex cellulase-xylanase system of Clostridium papyrosolvens. Also, progress toward completing the research of more recent projects is briefly summarized. Major accomplishments include: 1. Characterization of Clostridium phytofermentans, a cellulose-fermenting, ethanol-producing bacterium from forest soil. The characterization of a new cellulolytic species isolated from a cellulose-decomposing microbial consortium from forest soil was completed. This bacterium is remarkable for the high concentrations of ethanol produced during cellulose fermentation, typically more than twice the concentration produced by other species of cellulolytic clostridia. 2. Examination of the use of chitin as a source of carbon and nitrogen by cellulolytic microbes. We discovered that many cellulolytic anaerobes and facultative aerobes are able to use chitin as a source of both carbon and nitrogen. This major discovery expands our understanding of the biology of cellulose-fermenting bacteria and may lead to new applications for these microbes. 3. Comparative studies of the cellulase and chitinase systems of Cellulomonas uda. Results of these studies indicate that the chitinase and cellulase systems of this bacterium are distinct in terms of the proteins involved and the regulation of their production. 4. Characterization of the chitinase system of C. uda. A 70,000-Mr endochitinase, designated ChiA, was purified from C. uda culture supernatant fluids and characterized. 5. Analysis of chiA, which codes for the major enzymatic component of the chitinase system of C. uda. The gene encoding the endochitinase ChiA in C. uda was cloned, its complete nucleotide sequence was determined and its implications were investigated. 6. Formation of biofilms by C. uda on cellulose and chitin. Microscopic observations indicated that, under conditions of nitrogen limitation, C. uda cells grew as a biofilm attached tightly to the surface of cellulose or chitin. 7. Development of tools for a genetic approach to studies of cellulose fermentation by cellulolytic clostridia. We have explored the potential of various techniques, and obtained evidence indicating that Tn916 mutagenesis may be particularly effective in this regard. As part of this research, we identified the presence of a plasmid in one strain, which was cloned, sequenced, and analyzed for its utility in the development of vectors for genetic studies. 8. Effects of humic substances on cellulose degradation by anaerobic cellulolytic microbes. We determined that humic substances play an important role in the anaerobic cellulose decomposition and in the physiology of cellulose-fermenting soil bacteria. 9. Nitrogenases of cellulolytic clostridia. We described a nitrogenase gene from a cellulolytic clostridium and presented evidence, based on sequence analyses and conserved gene order, for lateral gene transfer between this bacterium and a methanogenic archaeon. 10. Characterization of Clostridium hungatei, a new N2-fixing cellulolytic species isolated from a methanogenic consortium from soil. 11. Understanding the molecular architecture of the multicomplex cellulase-xylanase system of Clostridium papyrosolvens. We discovered that C. papyrosolvens produces a multiprotein, multicom

Leschine, Susan

2009-10-31

349

Preparation of chitin butyrate by using phosphoryl mixed anhydride system.  

PubMed

Acylation of chitin with butyric acid was performed in the presence of trifluoroacetic anhydride/phosphoric acid mediated system. The products were characterized by (1)H NMR and FT-IR spectroscopy and their solubility was tested in different organic solvents. Inclusion of butyric acid moieties into the parent molecule was confirmed from the (1)H NMR and FT-IR spectra. FT-IR analysis revealed that the degree of acid substitution (DS) of the products was in a range of 1.9-2.38, which increased with increasing the amounts of butyric acid added to the reaction system. Degree of N-deacetylation (DD) of the products, as determined by (1)H NMR was between 54.2% and 65.6%. The products with DS >2.0 were soluble in dimethyl sulfoxide, N,N-dimethylformamide, tetrahydrofuran, methanol, acetone, chloroform, and acetic acid. PMID:21353204

Bhatt, Lok Ranjan; Kim, Bo Mi; Hyun, Kim; Kang, Kyung Hee; Lu, Chichong; Chai, Kyu Yun

2011-04-01

350

Multifaceted chitin/poly(lactic-co-glycolic) acid composite nanogels.  

PubMed

Cyto-compatible, 80nm sized chitin/PLGA composite nanogels (chit/PLGA-comp NGs) were prepared by regeneration method and characterized. The multifaceted chit/PLGA-comp NGs were surface modified with Au, Fe3O4, CdTe/ZnTe-QDs and umbelliferone, respectively. 185nm sized Au-chit/PLGA-comp NGs, 170nm sized QD-chit/PLGA-comp-NGs and 160nm sized Fe3O4-chit/PLGA-comp-NGs showed RF heating. The QD-chit/PLGA-comp-NGs and 180nm sized umb-chit/PLGA-comp-NGs were well uptaken by Escherichia coli, Staphylococcus aureus and Candida albicans. The chit/PLGA-comp NGs could be useful for microbial monitoring and RF application for cancer therapy. The preliminary data showed that multifaceted chit/PLGA-comp-NGs could be useful for hyperthermia for cancer treatment and microbial labelling and imaging. PMID:24685461

Rejinold, N Sanoj; Biswas, Raja; Chellan, Gopi; Jayakumar, R

2014-06-01

351

CHITIN TRANSFORMATION AND PESTICIDE INTERACTIONS IN A SIMULATED AQUATIC MICROENVIRONMENTAL SYSTEM  

EPA Science Inventory

Interactions between the structural animo-polysaccharide, chitin, and the organophosphate pesticide, azinphosmethyl (Guthion), have been studied in a controlled continuous flow-through microcosm. Pesticide-induced microbial population changes and increases in substrate utilizatio...

352

MOLECULAR TRACERS FOR SMOKE FROM CHARRING/BURNING OF CHITIN BIOPOLYMER. (R823990)  

EPA Science Inventory

Abstract Monosaccharide derivatives from the breakdown of cellulose are the major organic components of smoke particles emitted to the atmosphere from biomass burning. In urban areas a related biopolymer, chitin, may contribute markers to smoke from grilling/charring o...

353

Application of Spectroscopic Methods for Structural Analysis of Chitin and Chitosan  

PubMed Central

Chitin, the second most important natural polymer in the world, and its N-deacetylated derivative chitosan, have been identified as versatile biopolymers for a broad range of applications in medicine, agriculture and the food industry. Two of the main reasons for this are firstly the unique chemical, physicochemical and biological properties of chitin and chitosan, and secondly the unlimited supply of raw materials for their production. These polymers exhibit widely differing physicochemical properties depending on the chitin source and the conditions of chitosan production. The presence of reactive functional groups as well as the polysaccharide nature of these biopolymers enables them to undergo diverse chemical modifications. A complete chemical and physicochemical characterization of chitin, chitosan and their derivatives is not possible without using spectroscopic techniques. This review focuses on the application of spectroscopic methods for the structural analysis of these compounds. PMID:20559489

Kumirska, Jolanta; Czerwicka, Ma?gorzata; Kaczy?ski, Zbigniew; Bychowska, Anna; Brzozowski, Krzysztof; Thöming, Jorg; Stepnowski, Piotr

2010-01-01

354

Structural Investigations of Chitin and Chitosan Complexed with Iron or Tin  

NASA Astrophysics Data System (ADS)

Chitin (N-acetyl-glucosamine) and its derivative chitosan (glucosamine) bind with most transition and main group metals, including iron and tin. Using 57Fe and 119Sn Mössbauer Spectroscopy it is determined that an oxidation reaction occurs during the metal uptake. Data also supports a structure with more than one metal bonding site and shows the ability of the chitin and chitosan polymers to bind large concentrations of iron.

Gamblin, B. E.; Stevens, J. G.; Wilson, K. L.

1998-12-01

355

Preparation and Characterization of a Novel Co-processed Excipient of Chitin and Crystalline Mannitol  

Microsoft Academic Search

A co-processed excipient was prepared from commercially available crystalline mannitol and ?-chitin using direct compression\\u000a as well as spray, wet, and dry granulation. The effect of the ratio of the two components, percentage of lubricant and particle\\u000a size, on the properties of the prepared co-processed excipient has been investigated. ?-Chitin forms non-hygroscopic, highly\\u000a compactable, disintegrable compacts when co-processed with crystalline

Nidal Daraghmeh; Iyad Rashid; Mahmoud M. H. Al Omari; Stephen A. Leharne; Babur Z. Chowdhry; Adnan Badwan

2010-01-01

356

Hydrogen and oxygen in brine shrimp chitin reflect environmental water and dietary isotopic composition  

NASA Astrophysics Data System (ADS)

Hydrogen and oxygen isotope ratios of the common structural biopolymer chitin are a potential recorder of ecological and environmental information, but our understanding of the mechanisms of incorporation of H and O from environmental substrates into chitin is limited. We report the results of a set of experiments in which the isotopic compositions of environmental water and diet were varied independently in order to assess the contribution of these variables to the H and O isotopic composition of Artemia franciscana chitin. Hydrogen isotope ratios of chitin were strongly linearly correlated with both food and water, with approximately 26% of the hydrogen signal reflecting food and approximately 38% reflecting water. Oxygen isotopes were also strongly correlated with the isotopic composition of water and food, but whereas 69% of oxygen in chitin exchanged with environmental water, only 10% was derived from food. We propose that these observations reflect the position-specific, partial exchange of H and O atoms with brine shrimp body water during the processes of digestion and chitin biosynthesis. Comparison of culture experiments with a set of natural samples collected from the Great Salt Lake, UT in 2006 shows that, with some exceptions, oxygen isotope compositions of chitin track those of water, whereas hydrogen isotopes vary inversely with those of lake water. The different behavior of the two isotopic systems can be explained in terms of a dietary shift from allochthonous particulate matter with relatively higher ? 2H values in the early spring to autochthonous particulate matter with significantly lower ? 2H values in the late summer to autumn. These results suggest oxygen in chitin may be a valuable proxy for the oxygen isotopic composition of environmental water, whereas hydrogen isotope values from the same molecule may reveal ecological and biogeochemical changes within lakes.

Nielson, Kristine E.; Bowen, Gabriel J.

2010-03-01

357

Some chitin\\/chitosan derivatives for corrosion protection and waste water treatments  

Microsoft Academic Search

Chitin was extracted from locally collected shrimp shells. Chitosan was produced by alkali deacetylation of chitin. Poly(DEAEMA)-chitosan-graft-copolymer, poly(COOH)-chitosan-graft-copolymer, poly(V-OH)-chitosan-graft-copolymer, and carboxymethyl-chitosan were prepared. The extent of the preparation reactions was expressed as nitrogen content, carboxylic content and graft yield. The ability of the prepared compounds to adsorb heavy metals ions and some dyestuffs was studied. The prepared compounds were also

Sanaa M. El-Sawy; Yosreya M. Abu-Ayana; Fikry A. Abdel-Mohdy

2001-01-01

358

Chitin, Chitosan, and Glycated Chitosan Regulate Immune Responses: The Novel Adjuvants for Cancer Vaccine  

PubMed Central

With the development of cancer immunotherapy, cancer vaccine has become a novel modality for cancer treatment, and the important role of adjuvant has been realized recently. Chitin, chitosan, and their derivatives have shown their advantages as adjuvants for cancer vaccine. In this paper, the adjuvant properties of chitin and chitosan were discussed, and some detailed information about glycated chitosan and chitosan nanoparticles was also presented to illustrate the trend for future development. PMID:23533454

Li, Xiaosong; Min, Min; Du, Nan; Gu, Ying; Hode, Tomas; Naylor, Mark; Chen, Dianjun; Nordquist, Robert E.; Chen, Wei R.

2013-01-01

359

Methyl jasmonate induces expression of a novel Brassica juncea chitinase with two chitin-binding domains  

Microsoft Academic Search

We have cloned a 1.3 kb Brassica juncea cDNA encoding BjCHI1, a novel acidic chitinase with two chitin-binding domains that shows 62% identity to Nicotiana tabacum Chia1 chitinase. BjCHI1 is structurally unlike Chia1 that has one chitin-binding domain, but resembles Chia5 chitinase UDA1, the precursor of Urtica dioica agglutinin; however there is only 36.9% identity between them. We propose that

Kai-Jun Zhao; Mee-Len Chye

1999-01-01

360

Structural characterization of chitin and chitosan obtained by biological and chemical methods.  

PubMed

Chitin production was biologically achieved by lactic acid fermentation (LAF) of shrimp waste (Litopenaeus vannameii) in a packed bed column reactor with maximal percentages of demineralization (D(MIN)) and deproteinization (D(PROT)) after 96 h of 92 and 94%, respectively. This procedure also afforded high free astaxanthin recovery with up to 2400 ?g per gram of silage. Chitin product was also obtained from the shrimp waste by a chemical method using acid and alkali for comparison. The biologically obtained chitin (BIO-C) showed higher M(w) (1200 kDa) and crystallinity index (I(CR)) (86%) than the chemically extracted chitin (CH-C). A multistep freeze-pump-thaw (FPT) methodology was applied to obtain medium M(w) chitosan (400 kDa) with degree of acetylation (DA) ca. 10% from BIO-C, which was higher than that from CH-C. Additionally, I(CR) values showed the preservation of crystalline chitin structure in BIO-C derivatives at low DA (40-25%). Moreover, the FPT deacetylation of the attained BIO-C produced chitosans with bloc copolymer structure inherited from a coarse chitin crystalline morphology. Therefore, our LAF method combined with FPT proved to be an affective biological method to avoid excessive depolymerization and loss of crystallinity during chitosan production, which offers new perspective applications for this material. PMID:21790136

Pacheco, Neith; Garnica-Gonzalez, Mónica; Gimeno, Miquel; Bárzana, Eduardo; Trombotto, Stéphane; David, Laurent; Shirai, Keiko

2011-09-12

361

Facile route to produce chitin nanofibers as precursors for flexible and transparent gas barrier materials.  

PubMed

Chitin is the second most abundant biopolymer in nature and has tremendous potential in renewable materials for packaging, energy storage, reinforced composites, and biomedical engineering. Despite attractive properties, including biodegradability, antibacterial activity, and high strength, chitin is not utilized widely due to strong molecular interactions, which make solubilization and processing difficult. We report a high pressure homogenization route to produce pure chitin nanofibers (ChNFs) starting with a mildly acidic aqueous dispersion of purified crab ?-chitin. The well-dispersed ChNFs with diameter ?20 nm do not form strong network structures under conditions explored herein and can be directly processed into useful materials, bypassing the need to dissolve the chitin. Dried ChNFs form pure self-standing chitin films with the lowest to-date reported O2 and CO2 permeabilities of 0.006 and 0.018 barrer, respectively. Combined with high flexibility and optical transparency, these materials are ideal candidates for sustainable barrier packaging. PMID:25483821

Wu, Jie; Zhang, Kuang; Girouard, Natalie; Meredith, J Carson

2014-12-01

362

A pathway of chitosan formation in Mucor rouxii. Enzymatic deacetylation of chitin.  

PubMed

1. An enzyme that catalyzes hydrolysis of acetamido groups of chitin derivatives was found in the supernatant fraction of Mucor rouxii. 2. Partially O-hydroxyethylated chitin (glycol chitin) was used as a substrate in the purification and characterization of this enzyme. A 140-fold purification was obtained by means of ammonium sulfate fractionation followed by chromatography on carboxymethylcellulose and DEAE-cellulose. 3. The enzyme releases about 30% of the acetyl groups of glycol chitin, giving a product with a decreased sensitivity to lysozyme. The enzyme also deacetylates chitin and N-acetylchitooligoses, whereas it is inactive toward bacterial cell wall peptidoglycan, N-acetylated heparin, a polymer of N-acetylgalactosamine, di-N-acetylchitobiose and monomeric N-acetylglucosamine derivatives. 4. This enzyme shows a pH optimum of 5.5. The Km value for glycol chitin is 0.87 g/l or 2.6 mM with respect to monosaccharide residues. 5. The occurrence of this enzyme accounts for the formation of chitosan in fungi. PMID:240696

Araki, Y; Ito, E

1975-06-16

363

Molecular directionality in crystalline ?-chitin: hydrolysis by chitinases A and B from Serratia marcescens 2170  

PubMed Central

?-Chitin microfibrils were treated with ChiA and ChiB (chitinases A and B respectively) from Serratia marcescens 2170. The ?-chitin microfibrils were shortened, and the tips appeared narrowed and sharpened at both ends, after either consecutive or simultaneous degradation by ChiA and ChiB. Increased production of reducing sugars by simultaneous degradation (by ChiA and ChiB) of ?-chitin, but not of glycol chitin, suggests synergistic interactions between the two enzymes. A combined analysis using the tilt microdiffraction method to determine the crystallographic axes, together with the biotin–streptavidin–gold-labelling method specific to the reducing ends, was used to investigate the polarity of the degraded ?-chitin microcrystals. The digestion of the ?-chitin fibrils by ChiA occurred from the reducing end to the nonreducing end, whereas digestion by ChiB occurred from the non-reducing end to the reducing end. The results are in agreement with the previously determined three-dimensional structures of these enzymes. PMID:15717865

Hult, Eva-Lena; Katouno, Fuminori; Uchiyama, Taku; Watanabe, Takeshi; Sugiyama, Junji

2005-01-01

364

Fabrication of chitin microspheres and their multipurpose application as catalyst support and adsorbent.  

PubMed

In this study, novel chitin microspheres (CM) with diameters of 1010?m, 750?m, 490?m, 280?m were fabricated by employing the sol-gel transition method. Then the chitin microspheres served as the enabling platform for a range of applications including recyclable catalyst support and adsorbent. First, the freeze dried porous chitin microspheres were coated with dopamine to enhance the loading efficiency of a model biomacromolecule, ?-amylase. The immobilized enzyme (49.6mg/g) retained more than 95% of relative activity after 10 repeated cycles and exhibited easy recovery ability. Then porous magnetic chitin microspheres could be prepared, and the swollen porous polymer successfully controlled the growth of gold nanoparticles. The chitin/Au nanocomposite microspheres were a good recyclable catalyst due to the porous structure and a reduced dimension of the metal particles (r?5nm). Finally, the magnetic chitin microspheres were modified into an adsorbent for enhanced removal of a typical cationic compound, methylene blue from aqueous solution. PMID:25662687

Wang, Yuntao; Li, Yan; Liu, Shilin; Li, Bin

2015-04-20

365

Chitin Binding Proteins Act Synergistically with Chitinases in Serratia proteamaculans 568  

PubMed Central

Genome sequence of Serratia proteamaculans 568 revealed the presence of three family 33 chitin binding proteins (CBPs). The three Sp CBPs (Sp CBP21, Sp CBP28 and Sp CBP50) were heterologously expressed and purified. Sp CBP21 and Sp CBP50 showed binding preference to ?-chitin, while Sp CBP28 did not bind to chitin and cellulose substrates. Both Sp CBP21 and Sp CBP50 were synergistic with four chitinases from S. proteamaculans 568 (Sp ChiA, Sp ChiB, Sp ChiC and Sp ChiD) in degradation of ?- and ?-chitin, especially in the presence of external electron donor (reduced glutathione). Sp ChiD benefited most from Sp CBP21 or Sp CBP50 on ?-chitin, while Sp ChiB and Sp ChiD had major advantage with these Sp CBPs on ?-chitin. Dose responsive studies indicated that both the Sp CBPs exhibit synergism ?0.2 µM. The addition of both Sp CBP21 and Sp CBP50 in different ratios to a synergistic mixture did not significantly increase the activity. Highly conserved polar residues, important in binding and activity of CBP21 from S. marcescens (Sm CBP21), were present in Sp CBP21 and Sp CBP50, while Sp CBP28 had only one such polar residue. The inability of Sp CBP28 to bind to the test substrates could be attributed to the absence of important polar residues. PMID:22590591

Purushotham, Pallinti; Arun, P. V. Parvati Sai; Prakash, Jogadhenu S. S.; Podile, Appa Rao

2012-01-01

366

Catalytic Efficiency of Chitinase-D on Insoluble Chitinous Substrates Was Improved by Fusing Auxiliary Domains  

PubMed Central

Chitin is an abundant renewable polysaccharide, next only to cellulose. Chitinases are important for effective utilization of this biopolymer. Chitinase D from Serratia proteamaculans (SpChiD) is a single domain chitinase with both hydrolytic and transglycosylation (TG) activities. SpChiD had less of hydrolytic activity on insoluble polymeric chitin substrates due to the absence of auxiliary binding domains. We improved catalytic efficiency of SpChiD in degradation of insoluble chitin substrates by fusing with auxiliary domains like polycystic kidney disease (PKD) domain and chitin binding protein 21 (CBP21). Of the six different SpChiD fusion chimeras, two C-terminal fusions viz. ChiD+PKD and ChiD+CBP resulted in improved hydrolytic activity on ?- and ?-chitin, respectively. Time-course degradation of colloidal chitin also confirmed that these two C-terminal SpChiD fusion chimeras were more active than other chimeras. More TG products were produced for a longer duration by the fusion chimeras ChiD+PKD and PKD+ChiD+CBP. PMID:25615694

Madhuprakash, Jogi; El Gueddari, Nour Eddine; Moerschbacher, Bruno M.; Podile, Appa Rao

2015-01-01

367

Preparation, characterization, and antimicrobial activity of chitin nanofibrils reinforced carrageenan nanocomposite films.  

PubMed

Present study illustrates the preparation of chitin nanofibrils (CNF) reinforced carrageenan nanocomposite films by the solution-casting technique. CNF was prepared by acid hydrolysis of chitin, followed by high speed homogenization and sonication. FTIR result demonstrated that the chemical structure of chitin had not changed after acid hydrolysis. However, the crystalinity of CNF was found to be higher than chitin. The crystallite size of chitin and CNF was 4.73 and 6.27nm, respectively. The char content at 600°C of chitin (19.2%) was lower than the CNF (25%). The carrageenan/CNF composite films were smooth and flexible and the CNF was dispersed uniformly in the carrageenan polymer matrix. The tensile strength and modulus of carrageenan film were increased significantly (p<0.05) after CNF reinforcement with up to 5 wt%, however, elongation at break, water vapor permeability, and transparency decreased slightly. Carrageenan/CNF nanocomposite films showed strong antibacterial activity against a Gram-positive food-borne pathogen, Listeria monocytogenes. PMID:25498660

Shankar, Shiv; Reddy, Jeevan Prasad; Rhim, Jong-Whan; Kim, Hee-Yun

2015-03-01

368

Surface characteristics of polyurethane elastomers based on chitin/1,4-butane diol blends.  

PubMed

Biodegradable polyurethane elastomers with tunable hydrophobicity were synthesized by step-growth polymerization techniques using poly(epsilon-caprolactone) (PCL) and 4,4'-diphenylmethane diisocyanate (MDI). The prepolymer was extended with different mass ratios of chitin and 1,4-butane diol (BDO). The effect of chitin contents in chain extenders (CE) proportion on surface properties was studied and investigated. Incorporation of chitin contents into the final PU showed decrease in surface free energy and its polar component. Simultaneously, the work of water adhesion to polymer decreases significantly by increasing the chitin contents in the synthesized polymer. Contact angle measurement, water absorption and swelling behavior of the synthesized polyurethane samples were affected by varying the chitin contents in the chemical composition of the final PU. The interactions of the final PU films with solvents on the surface were displayed clear dependent on the contents of chitin in to the final polyurethane formulation. The results of different tests demonstrated that the synthesized products are a potential candidate as non-absorbable suture as previously investigated into their in vitro biocompatibility and non-toxicity [K.M. Zia, M. Zuber, I.A. Bhatti, M. Barikani, M.A. Sheikh, Int. J. Biol. Macromol. 44 (2009) 18-22]. PMID:19133290

Zia, Khalid Mahmood; Barikani, Mehdi; Zuber, Mohammad; Bhatti, Ijaz Ahmad; Barmar, Mohammad

2009-03-01

369

Squid-Derived Chitin Oligosaccharides Are a Chemotactic Signal during Colonization by Vibrio fischeri  

PubMed Central

Chitin, a polymer of N-acetylglucosamine (GlcNAc), is noted as the second most abundant biopolymer in nature. Chitin serves many functions for marine bacteria in the family Vibrionaceae (“vibrios”), in some instances providing a physical attachment site, inducing natural genetic competence, and serving as an attractant for chemotaxis. The marine luminous bacterium Vibrio fischeri is the specific symbiont in the light-emitting organ of the Hawaiian bobtail squid, Euprymna scolopes. The bacterium provides the squid with luminescence that the animal uses in an antipredatory defense, while the squid supports the symbiont's nutritional requirements. V. fischeri cells are harvested from seawater during each host generation, and V. fischeri is the only species that can complete this process in nature. Furthermore, chitin is located in squid hemocytes and plays a nutritional role in the symbiosis. We demonstrate here that chitin oligosaccharides produced by the squid host serve as a chemotactic signal for colonizing bacteria. V. fischeri uses the gradient of host chitin to enter the squid light organ duct and colonize the animal. We provide evidence that chitin serves a novel function in an animal-bacterial mutualism, as an animal-produced bacterium-attracting synomone. PMID:22522684

Schaefer, Amy L.; Brennan, Caitlin A.; Heath-Heckman, Elizabeth A. C.; DeLoney-Marino, Cindy R.; McFall-Ngai, Margaret J.

2012-01-01

370

Evaluation of biocompatibility and mechanical behavior of polyurethane elastomers based on chitin/1,4-butane diol blends.  

PubMed

Chitin based polyurethane elastomers with potential as biomedical implants with tunable mechanical properties were synthesized by step growth polymerization techniques using poly(epsilon-caprolactone) (PCL) and 4,4'-diphenylmethane diisocyanate (MDI). The prepolymer was extended with different mass ratios of chitin and 1,4-butane diol (BDO). Molecular characterization was done using FTIR, 1H NMR and 13C NMR techniques. The mechanical properties of these polymers were improved with increase in the chitin contents. Optimum mechanical properties were obtained from elastomers extended with chitin in comparison to elastomers extended with BDO. Cytotoxicity of the synthesized polyurethane samples was affected by varying the chitin contents in the chemical composition of the final polyurethane (PU). It is revealed that the final polymers extended with chitin are preferred candidates for surgical threads with on going investigations into their in vitro biocompatibility and non-toxicity. PMID:18930759

Zia, Khalid Mahmood; Zuber, Mohammad; Bhatti, Ijaz Ahmad; Barikani, Mehdi; Sheikh, Munir Ahmad

2009-01-01

371

Targeting glycogen synthase kinase-3 in insulin signalling.  

PubMed

The renewed interest in an enzyme first discovered over 25 years ago stems from the potential of inhibitors of this enzyme to treat conditions as diverse as diabetes, Alzheimer's disease, stroke and bipolar disorder, and even to enhance the repopulating capacity of transplanted haematopoietic stem cells. The emergence of the first few potent and specific glycogen synthase kinase-3 (GSK-3) inhibitors will end years of speculation on their potential and finally allow the impact of GSK-3 inhibitors to be evaluated clinically. The next few years are likely to be particularly exciting ones for fans of this old enzyme. This review focuses on the role of GSK-3 in the insulin signalling pathway and highlights the evidence implicating the enzyme in insulin resistance. Pharmacological in vitro and in vivo proof-of-concept studies are also discussed, which establish the therapeutic potential of GSK-3 inhibitors as agents for the treatment of Type 2 diabetes. PMID:16706683

Frame, Sheelagh; Zheleva, Daniella

2006-06-01

372

Oral Administration of Chitin Down-Regulates Serum IgE Levels and Lung Eosinophilia in the Allergic Mouse1  

Microsoft Academic Search

Previous studies showed that local macrophages phagocytose nonantigenic chitin particles (1-10 mm polymers of N-acetyl-D- glucosamine) through mannose receptors and produce IL-12, IL-18, and TNF-a. These cytokines lead to the production of IFN-g by NK cells. To determine whether chitin could down-regulate Th2 responses, chitin was given orally (8 mg\\/day for 3 days before and 13 days during ragweed allergen

Yoshimi Shibata; L. Ann Foster; John F. Bradfield; Quentin N. Myrvik

373

Determination of the degree of acetylation of chitin materials by 13C CP\\/MAS NMR spectroscopy  

Microsoft Academic Search

13C CP\\/MAS NMR spectroscopy has been shown to be a powerful tool to quantify the degree of acetylation of chitin and chitosan. In order to optimise the parameters which afford quantitative 13C cross-polarisation magic-angle spinning NMR spectra, a detailed relaxation study has been carried out on selected chitin and deacetylated chitin samples. A relaxation delay of 5 s and a

M. L. Duarte; M. C. Ferreira; M. R. Marvão; João Rocha

2001-01-01

374

Three-dimensional chitin-based scaffolds from Verongida sponges (Demospongiae: Porifera). Part II: Biomimetic potential and applications.  

PubMed

In order to evaluate the biomedical potential of three-dimensional chitinous scaffolds of poriferan origin, chondrocyte culturing experiments were performed. It was shown for the first time that freshly isolated chondrocytes attached well to the chitin scaffold and synthesized an extracellular matrix similar to that found in other cartilage tissue engineering constructs. Chitin scaffolds also supported deposition of a proteoglycan-rich extracellular matrix of chondrocytes seeded bioconstructs in an in vivo environment. We suggest that chitin sponge scaffolds, apart from the demonstrated biomedical applications, are highly optimized structures for use as filtering systems, templates for biomineralization as well as metallization in order to produce catalysts. PMID:20478334

Ehrlich, H; Steck, E; Ilan, M; Maldonado, M; Muricy, G; Bavestrello, G; Kljajic, Z; Carballo, J L; Schiaparelli, S; Ereskovsky, A; Schupp, P; Born, R; Worch, H; Bazhenov, V V; Kurek, D; Varlamov, V; Vyalikh, D; Kummer, K; Sivkov, V V; Molodtsov, S L; Meissner, H; Richter, G; Hunoldt, S; Kammer, M; Paasch, S; Krasokhin, V; Patzke, G; Brunner, E; Richter, W

2010-08-01

375

Chitin amendment increases soil suppressiveness toward plant pathogens and modulates the actinobacterial and oxalobacteraceal communities in an experimental agricultural field.  

PubMed

A long-term experiment on the effect of chitin addition to soil on the suppression of soilborne pathogens was set up and monitored for 8 years in an experimental field, Vredepeel, The Netherlands. Chitinous matter obtained from shrimps was added to soil top layers on two different occasions, and the suppressiveness of soil toward Verticillium dahliae, as well as plant-pathogenic nematodes, was assessed, in addition to analyses of the abundances and community structures of members of the soil microbiota. The data revealed that chitin amendment had raised the suppressiveness of soil, in particular toward Verticillium dahliae, 9 months after the (second) treatment, extending to 2 years following treatment. Moreover, major effects of the added chitin on the soil microbial communities were detected. First, shifts in both the abundances and structures of the chitin-treated soil microbial communities, both of total soil bacteria and fungi, were found. In addition, the abundances and structures of soil actinobacteria and the Oxalobacteraceae were affected by chitin. At the functional gene level, the abundance of specific (family-18 glycoside hydrolase) chitinase genes carried by the soil bacteria also revealed upshifts as a result of the added chitin. The effects of chitin noted for the Oxalobacteraceae were specifically related to significant upshifts in the abundances of the species Duganella violaceinigra and Massilia plicata. These effects of chitin persisted over the time of the experiment. PMID:23811512

Cretoiu, Mariana Silvia; Korthals, Gerard W; Visser, Johnny H M; van Elsas, Jan Dirk

2013-09-01

376

Squalene synthase inhibition: a novel target for the management of dyslipidemia.  

PubMed

A new class of compounds, known as squalene synthase inhibitors, has recently reached phase III clinical trials and may provide another therapeutic option for clinicians to improve risk management of low-density lipoprotein cholesterol (LDL-C). The clinical need for another LDL-C-lowering therapy is evident by the inability to achieve an LDL-C target of less than 70 mg/dL in the majority of very high-risk patients on statin monotherapy. Human clinical trial data with TAK-475, a novel and potent inhibitor of squalene synthase, have not yet been published. PMID:17169251

Davidson, Michael H

2007-01-01

377

Design, synthesis and biological evaluation of N-alkyl or aryl substituted isoindigo derivatives as potential dual cyclin-dependent kinase 2 (CDK2)/glycogen synthase kinase 3? (GSK-3?) phosphorylation inhibitors.  

PubMed

A series of N-alkyl or aryl substituted isoindigo derivatives have been synthesized and their anti-proliferative activity was evaluated by Sulforhodamine B (SRB) assay. Some of the target compounds exhibited significant antitumor activity, including compounds 6h and 6k (against K562 cells), 6i (against HeLa cells) and 6j (against A549 cells). N-(p-methoxy-phenyl)-isoindigo (6k) exhibited a high and selective anti-proliferative activity against K562 cells (IC50 7.8 ?M) and induced the apoptosis of K562 cells in a dose-dependent manner. Compound 6k arrested the cell cycle at S phase in K562 cells by decreasing the expression of cyclin A and CDK2, which played critical roles in DNA replication and passage through G2 phase. Moreover, compound 6k down-regulated the expression of p-GSK-3? (Ser9), ?-catenin and c-myc proteins, up-regulated the expression of GSK-3?, consequently, suppressed Wnt/?-catenin signaling pathway and induced the apoptosis of K562 cells. The binding mode of compound 6k with GSK-3? was simulated using molecular docking tools. All of these studies gave a better understanding to the molecular mechanisms of this class of agents and clues to develop dual CDK2/GSK-3? (Ser9) phosphorylation inhibitors applied in cancer chemotherapy. PMID:25151579

Zhao, Ping; Li, Yanzhong; Gao, Guangwei; Wang, Shuai; Yan, Yun; Zhan, Xiaoping; Liu, Zenglu; Mao, Zhenmin; Chen, Shaoxiong; Wang, Liqun

2014-10-30

378

Squid Pen Chitin Chitooligomers as Food Colorants Absorbers  

PubMed Central

One of the most promising applications of chitosanase is the conversion of chitinous biowaste into bioactive chitooligomers (COS). TKU033 chitosanase was induced from squid pen powder (SPP)-containing Bacillus cereus TKU033 medium and purified by ammonium sulfate precipitation and column chromatography. The enzyme was relatively more thermostable in the presence of the substrate and had an activity of 93% at 50 °C in a pH 5 buffer solution for 60 min. Furthermore, the enzyme used for the COS preparation was also studied. The enzyme products revealed various mixtures of COS that with different degrees of polymerization (DP), ranging from three to nine. In the culture medium, the fermented SPP was recovered, and it displayed a better adsorption rate (up to 96%) for the disperse dyes than the water-soluble food colorants, Allura Red AC (R40) and Tartrazne (Y4). Fourier transform-infrared spectroscopic (FT-IR) analysis proved that the adsorption of the dyes onto fermented SPP was a physical adsorption. Results also showed that fermented SPP was a favorable adsorber and could be employed as low-cost alternative for dye removal in wastewater treatment. PMID:25608726

Liang, Tzu-Wen; Huang, Chih-Ting; Dzung, Nguyen Anh; Wang, San-Lang

2015-01-01

379

Chitin butyrate coated electrospun nylon-6 fibers for biomedical applications  

NASA Astrophysics Data System (ADS)

In this study, we describe the preparation and characterizations of chitin butyrate (CB) coated nylon-6 nanofibers using single-spinneret electrospinning of blends solution. The physicochemical properties of nylon-6 composite fibers with different proportions of CB to nylon-6 were determined using FE-SEM, TEM, FT-IR spectroscopy, and water contact angle measurement. FE-SEM and TEM images revealed that the nylon-6 and CB were immiscible in the as-spun nanofibers, and phase separated nanofiber morphology becomes more pronounced with increasing amounts of CB. The bone formation ability of composite fibers was evaluated by incubating in biomimetic simulated body fluid. In order to assay the cytocompatibility and cell behavior on the composite scaffolds, osteoblast cells were seeded on the matrix. Results suggest that the deposition of CB layer on the surface of nylon-6 could increase its cell compatibility and bone formation ability. Therefore, as-synthesized nanocomposite fibrous mat has great potentiality in hard tissue engineering.

Pant, Hem Raj; Kim, Han Joo; Bhatt, Lok Ranjan; Joshi, Mahesh Kumar; Kim, Eun Kyo; Kim, Jeong In; Abdal-hay, Abdalla; Hui, K. S.; Kim, Cheol Sang

2013-11-01

380

Squid pen chitin chitooligomers as food colorants absorbers.  

PubMed

One of the most promising applications of chitosanase is the conversion of chitinous biowaste into bioactive chitooligomers (COS). TKU033 chitosanase was induced from squid pen powder (SPP)-containing Bacillus cereus TKU033 medium and purified by ammonium sulfate precipitation and column chromatography. The enzyme was relatively more thermostable in the presence of the substrate and had an activity of 93% at 50 °C in a pH 5 buffer solution for 60 min. Furthermore, the enzyme used for the COS preparation was also studied. The enzyme products revealed various mixtures of COS that with different degrees of polymerization (DP), ranging from three to nine. In the culture medium, the fermented SPP was recovered, and it displayed a better adsorption rate (up to 96%) for the disperse dyes than the water-soluble food colorants, Allura Red AC (R40) and Tartrazne (Y4). Fourier transform-infrared spectroscopic (FT-IR) analysis proved that the adsorption of the dyes onto fermented SPP was a physical adsorption. Results also showed that fermented SPP was a favorable adsorber and could be employed as low-cost alternative for dye removal in wastewater treatment. PMID:25608726

Liang, Tzu-Wen; Huang, Chih-Ting; Dzung, Nguyen Anh; Wang, San-Lang

2015-01-01

381

Modification of chitin with kraft lignin and development of new biosorbents for removal of cadmium(II) and nickel(II) ions.  

PubMed

Novel, functional materials based on chitin of marine origin and lignin were prepared. The synthesized materials were subjected to physicochemical, dispersive-morphological and electrokinetic analysis. The results confirm the effectiveness of the proposed method of synthesis of functional chitin/lignin materials. Mechanism of chitin modification by lignin is based on formation of hydrogen bonds between chitin and lignin. Additionally, the chitin/lignin materials were studied from the perspective of waste water treatment. The synthetic method presented in this work shows an attractive and facile route for producing low-cost chitin/lignin biosorbents with high efficiency of nickel and cadmium adsorption (88.0% and 98.4%, respectively). The discovery of this facile method of synthesis of functional chitin/lignin materials will also have a significant impact on the problematic issue of the utilization of chitinous waste from the seafood industry, as well as lignin by-products from the pulp and paper industry. PMID:24727394

Wysokowski, Marcin; Klapiszewski, ?ukasz; Moszy?ski, Dariusz; Bartczak, Przemys?aw; Szatkowski, Tomasz; Majchrzak, Izabela; Siwi?ska-Stefa?ska, Katarzyna; Bazhenov, Vasilii V; Jesionowski, Teofil

2014-04-01

382

Modification of Chitin with Kraft Lignin and Development of New Biosorbents for Removal of Cadmium(II) and Nickel(II) Ions  

PubMed Central

Novel, functional materials based on chitin of marine origin and lignin were prepared. The synthesized materials were subjected to physicochemical, dispersive-morphological and electrokinetic analysis. The results confirm the effectiveness of the proposed method of synthesis of functional chitin/lignin materials. Mechanism of chitin modification by lignin is based on formation of hydrogen bonds between chitin and lignin. Additionally, the chitin/lignin materials were studied from the perspective of waste water treatment. The synthetic method presented in this work shows an attractive and facile route for producing low-cost chitin/lignin biosorbents with high efficiency of nickel and cadmium adsorption (88.0% and 98.4%, respectively). The discovery of this facile method of synthesis of functional chitin/lignin materials will also have a significant impact on the problematic issue of the utilization of chitinous waste from the seafood industry, as well as lignin by-products from the pulp and paper industry. PMID:24727394

Wysokowski, Marcin; Klapiszewski, ?ukasz; Moszy?ski, Dariusz; Bartczak, Przemys?aw; Szatkowski, Tomasz; Majchrzak, Izabela; Siwi?ska-Stefa?ska, Katarzyna; Bazhenov, Vasilii V.; Jesionowski, Teofil

2014-01-01

383

Applying Molecular Dynamics Simulations to Identify Rarely Sampled Ligand-bound Conformational States of Undecaprenyl Pyrophosphate Synthase, an Antibacterial Target  

SciTech Connect

Undecaprenyl pyrophosphate synthase is a cis-prenyltransferase enzyme, which is required for cell wall biosynthesis in bacteria. Undecaprenyl pyrophosphate synthase is an attractive target for antimicrobial therapy. We performed long molecular dynamics simulations and docking studies on undecaprenyl pyrophosphate synthase to investigate its dynamic behavior and the influence of protein flexibility on the design of undecaprenyl pyrophosphate synthase inhibitors. We also describe the first X-ray crystallographic structure of Escherichia coli apo-undecaprenyl pyrophosphate synthase. The molecular dynamics simulations indicate that undecaprenyl pyrophosphate synthase is a highly flexible protein, with mobile binding pockets in the active site. By carrying out docking studies with experimentally validated undecaprenyl pyrophosphate synthase inhibitors using high- and low-populated conformational states extracted from the molecular dynamics simulations, we show that structurally dissimilar compounds can bind preferentially to different and rarely sampled conformational states. By performing structural analyses on the newly obtained apo-undecaprenyl pyrophosphate synthase and other crystal structures previously published, we show that the changes observed during the molecular dynamics simulation are very similar to those seen in the crystal structures obtained in the presence or absence of ligands. We believe that this is the first time that a rare 'expanded pocket' state, key to drug design and verified by crystallography, has been extracted from a molecular dynamics simulation.

Sinko, William; de Oliveira, César; Williams, Sarah; Van Wynsberghe, Adam; Durrant, Jacob D.; Cao, Rong; Oldfield, Eric; McCammon, J. Andrew (UIUC); (UCSD); (Hamilton)

2012-04-30

384

Applying Molecular Dynamics Simulations to Identify Rarely Sampled Ligand-bound Conformational States of Undecaprenyl Pyrophosphate Synthase, an Antibacterial Target  

PubMed Central

Undecaprenyl pyrophosphate synthase is a cis-prenyltransferase enzyme, which is required for cell wall biosynthesis in bacteria. Undecaprenyl pyrophosphate synthase is an attractive target for antimicrobial therapy. We performed long molecular dynamics simulations and docking studies on undecaprenyl pyrophosphate synthase to investigate its dynamic behavior and the influence of protein flexibility on the design of undecaprenyl pyrophosphate synthase inhibitors. We also describe the first X-ray crystallographic structure of Escherichia coli apo-undecaprenyl pyrophosphate synthase. The molecular dynamics simulations indicate that undecaprenyl pyrophosphate synthase is a highly flexible protein, with mobile binding pockets in the active site. By carrying out docking studies with experimentally validated undecaprenyl pyrophosphate synthase inhibitors using high- and low-populated conformational states extracted from the molecular dynamics simulations, we show that structurally dissimilar compounds can bind preferentially to different and rarely sampled conformational states. By performing structural analyses on the newly obtained apo-undecaprenyl pyrophosphate synthase and other crystal structures previously published, we show that the changes observed during the molecular dynamics simulation are very similar to those seen in the crystal structures obtained in the presence or absence of ligands. We believe that this is the first time that a rare ‘expanded pocket’ state, key to drug design and verified by crystallography, has been extracted from a molecular dynamics simulation. PMID:21294851

Sinko, William; de Oliveira, César; Williams, Sarah; Van Wynsberghe, Adam; Durrant, Jacob D; Cao, Rong; Oldfield, Eric; McCammon, J Andrew

2011-01-01

385

Chitin-based materials in tissue engineering: applications in soft tissue and epithelial organ.  

PubMed

Chitin-based materials and their derivatives are receiving increased attention in tissue engineering because of their unique and appealing biological properties. In this review, we summarize the biomedical potential of chitin-based materials, specifically focusing on chitosan, in tissue engineering approaches for epithelial and soft tissues. Both types of tissues play an important role in supporting anatomical structures and physiological functions. Because of the attractive features of chitin-based materials, many characteristics beneficial to tissue regeneration including the preservation of cellular phenotype, binding and enhancement of bioactive factors, control of gene expression, and synthesis and deposition of tissue-specific extracellular matrix are well-regulated by chitin-based scaffolds. These scaffolds can be used in repairing body surface linings, reconstructing tissue structures, regenerating connective tissue, and supporting nerve and vascular growth and connection. The novel use of these scaffolds in promoting the regeneration of various tissues originating from the epithelium and soft tissue demonstrates that these chitin-based materials have versatile properties and functionality and serve as promising substrates for a great number of future applications. PMID:21673932

Yang, Tsung-Lin

2011-01-01

386

Unexpected destruction of chitosomal chitin synthetase by an endogenous protease during sucrose density gradient purification.  

PubMed

Because of their intrinsic low buoyant density, chitosomes can be separated from crude cell homogenates (1000 g or 35,000 g supernatants) of Mucor rouxii by isopycnic sedimentation in sucrose density gradients. To accelerate and simplify the isolation of chitosomes, we centrifuged the cell-free extracts at ultrahigh speed (in a fixed-angle rotor at forces up to 311,000 g Rav) and found that the duration of centrifugation was critical. Prolonged centrifugation at ultrahigh speed caused severe distortion of the chitin synthetase profile in the gradient as the peak of chitosomal chitin synthetase nearly disappeared. We traced the problem to a soluble protease(s) that moved into the chitosome band during protracted centrifugation and destroyed the chitin synthetase activity. The interfering protease was a soluble protein with a sedimentation coefficient of 4.6 S and a pH optimum of 7-7.5, and it was sensitive to PMSF (phenylmethylsulfonyl fluoride), indicating that it was a serine protease. Unlike other proteases, it destroyed chitin synthetase but did not activate the chitin synthetase zymogen. The interfering protease could be eliminated either by adding PMSF to the cells immediately after breakage or by removing the upper part of the sucrose gradient midway through the centrifugation of the cell-free extract and then completing the sedimentation with the 'decapitated' gradient. PMID:1939371

Kamada, T; Bracker, C E; Lippman, E; Bartnicki-Garcia, S

1991-07-01

387

Characterization of a Chitin-Binding Protein from Bacillus thuringiensis HD-1  

PubMed Central

Strains of Bacillus thuringiensis produce insecticidal proteins. These strains have been isolated from diverse ecological niches, such as soil, phylloplane, insect cadavers and grain dust. To effectively propagate, these strains produce a range of molecules that facilitate its multiplication in a competing environment. In this report, we have examined synthesis of a chitin-binding protein and evaluated its effect on fungi encountered in environment and its interaction with insecticidal proteins synthesized by B. thuringiensis. The gene encoding chitin-binding protein has been cloned and expressed. The purified protein has been demonstrated to interact with Cry insecticidal protein, Cry1Ac by Circular Dichrosim spectroscopy (CD) and in vitro pull down assays. The chitin-binding protein potentiates insecticidal activity of bacillar insecticidal protein, Cry1Ac. Further, chitin-binding protein was fungistatic against several soil fungi. The chitin binding protein is expressed in spore mother cell and deposited along with insecticidal protein, Cry1Ac. It interacts with Cry1Ac to potentiate its insecticidal activity and facilitate propagation of Bacillus strain in environment by inhibiting growth of certain fungi. PMID:23824872

Gupta, Rani; Vimala, Y.; Bhatnagar, Raj K.

2013-01-01

388

A LysM receptor-like kinase plays a critical role in chitin signaling and fungal resistance in Arabidopsis  

Technology Transfer Automated Retrieval System (TEKTRAN)

Chitin, a polymer of N-acetyl-D-glucosamine, is found in fungal cell walls, but not in plants. Plant cells are capable of perceiving chitin fragments (chitooligosaccharides) to trigger plant defense. We identified a LysM receptor-like protein (AtLysM RLK1) that is required for the perception of chit...

389

Fungal effector Ecp6 outcompetes host immune receptor for chitin binding through intrachain LysM dimerization  

PubMed Central

While host immune receptors detect pathogen-associated molecular patterns to activate immunity, pathogens attempt to deregulate host immunity through secreted effectors. Fungi employ LysM effectors to prevent recognition of cell wall-derived chitin by host immune receptors, although the mechanism to compete for chitin binding remained unclear. Structural analysis of the LysM effector Ecp6 of the fungal tomato pathogen Cladosporium fulvum reveals a novel mechanism for chitin binding, mediated by intrachain LysM dimerization, leading to a chitin-binding groove that is deeply buried in the effector protein. This composite binding site involves two of the three LysMs of Ecp6 and mediates chitin binding with ultra-high (pM) affinity. Intriguingly, the remaining singular LysM domain of Ecp6 binds chitin with low micromolar affinity but can nevertheless still perturb chitin-triggered immunity. Conceivably, the perturbation by this LysM domain is not established through chitin sequestration but possibly through interference with the host immune receptor complex. DOI: http://dx.doi.org/10.7554/eLife.00790.001 PMID:23840930

Kombrink, Anja; Hansen, Guido; Valkenburg, Dirk-Jan

2013-01-01

390

A possible structural model of members of the CPF family of cuticular proteins implicating binding to components other than chitin  

Microsoft Academic Search

The physical properties of cuticle are determined by the structure of its two major components, cuticular proteins (CPs) and chitin, and, also, by their interactions.A common consensus region (extended R&R Consensus) found in the majority of cuticular proteins, the CPRs, binds to chitin. Previous work established that ?-pleated sheet predominates in the Consensus region and we proposed that it is

Nikos C. Papandreou; Vassiliki A. Iconomidou; Judith H. Willis; Stavros J. Hamodrakas

2010-01-01

391

A LysM Receptor-like Kinase Plays a Critical Role in Chitin Signaling and Fungal Resistance in Arabidopsis  

Technology Transfer Automated Retrieval System (TEKTRAN)

Chitin, a polymer of N-acetyl-D-glucosamine, is found in fungal cell walls, but not in plants. Plant cells are capable of perceiving chitin fragments (chitooligosaccharides) to trigger plant defense. We identified a LysM receptor-like protein (AtLysM RLK1) that is required for the perception of chit...

392

Multisite prenylation of 4-substituted tryptophans by dimethylallyltryptophan synthase.  

PubMed

The aromatic prenyltransferase dimethylallyltryptophan synthase in Claviceps purpurea catalyzes the normal prenylation of tryptophan at C4 of the indole nucleus in the first committed step of ergot alkaloid biosynthesis. 4-Methyltryptophan is a competitive inhibitor of the enzyme that has been used in kinetic studies. Upon investigation of background activity during incubations of 4-methyltryptophan with dimethylallyl diphosphate, we found that the analogue was an alternate substrate, which gave four products. The structures of three of these compounds were established by (1)H NMR and 2D NMR studies and revealed that dimethylallyltryptophan synthase catalyzed both normal and reverse prenylation at C3 of the indole ring and normal prenylation of N1. Similarly, 4-methoxytryptophan was an alternate substrate, giving normal prenylation at C5 as the major product. 4-Aminotryptophan, another alternate substrate, gave normal prenylation at C5 and C7. The ability of dimethylallyltryptophan synthase to prenylate at five different sites on the indole nucleus, with normal and reverse prenylation at one of the sites, is consistent with a dissociative electrophilic alkylation of the indole ring, where orientation of the substrates within the active site and substituent electronic effects determine the position and type of prenylation. These results suggest a common mechanism for prenylation of tryptophan by all of the members of the structurally related dimethylallyltryptophan synthase family. PMID:23301871

Rudolf, Jeffrey D; Wang, Hong; Poulter, C Dale

2013-02-01

393

Mechanism of action and inhibition of dehydrosqualene synthase  

PubMed Central

“Head-to-head” terpene synthases catalyze the first committed steps in sterol and carotenoid biosynthesis: the condensation of two isoprenoid diphosphates to form cyclopropylcarbinyl diphosphates, followed by ring opening. Here, we report the structures of Staphylococcus aureus dehydrosqualene synthase (CrtM) complexed with its reaction intermediate, presqualene diphosphate (PSPP), the dehydrosqualene (DHS) product, as well as a series of inhibitors. The results indicate that, on initial diphosphate loss, the primary carbocation so formed bends down into the interior of the protein to react with C2,3 double bond in the prenyl acceptor to form PSPP, with the lower two-thirds of both PSPP chains occupying essentially the same positions as found in the two farnesyl chains in the substrates. The second-half reaction is then initiated by the PSPP diphosphate returning back to the Mg2+ cluster for ionization, with the resultant DHS so formed being trapped in a surface pocket. This mechanism is supported by the observation that cationic inhibitors (of interest as antiinfectives) bind with their positive charge located in the same region as the cyclopropyl carbinyl group; that S-thiolo-diphosphates only inhibit when in the allylic site; activity results on 11 mutants show that both DXXXD conserved domains are essential for PSPP ionization; and the observation that head-to-tail isoprenoid synthases as well as terpene cyclases have ionization and alkene-donor sites which spatially overlap those found in CrtM. PMID:21098670

Lin, Fu-Yang; Liu, Chia-I; Liu, Yi-Liang; Zhang, Yonghui; Wang, Ke; Jeng, Wen-Yih; Ko, Tzu-Ping; Cao, Rong; Wang, Andrew H.-J.; Oldfield, Eric

2010-01-01

394

Effects of Chitin and Its Derivative Chitosan on Postharvest Decay of Fruits: A Review  

PubMed Central

Considerable economic losses to harvested fruits are caused by postharvest fungal decay during transportation and storage, which can be significantly controlled by synthetic fungicides. However, considering public concern over pesticide residues in food and the environment, there is a need for safer alternatives for the control of postharvest decay to substitute synthetic fungicides. As the second most abundant biopolymer renewable source in nature, chitin and its derivative chitosan are widely used in controlling postharvest decay of fruits. This review aims to introduce the effect of chitin and chitosan on postharvest decay in fruits and the possible modes of action involved. We found most of the actions discussed in these researches rest on physiological mechanisms. All of the mechanisms are summarized to lay the groundwork for further studies which should focus on the molecular mechanisms of chitin and chitosan in controlling postharvest decay of fruits. PMID:21541034

Zhang, Hongyin; Li, Renping; Liu, Weimin

2011-01-01

395

Bridging peripheral nerves using a deacetyl chitin conduit combined with short-term electrical stimulation  

PubMed Central

Previous studies have demonstrated that deacetyl chitin conduit nerve bridging or electrical stimulation can effectively promote the regeneration of the injured peripheral nerve. We hypothesized that the combination of these two approaches could result in enhanced regeneration. Rats with right sciatic nerve injury were subjected to deacetyl chitin conduit bridging combined with electrical stimulation (0.1 ms, 3 V, 20 Hz, for 1 hour). At 6 and 12 weeks after treatment, nerve conduction velocity, myelinated axon number, fiber diameter, axon diameter and the thickness of the myelin sheath in the stimulation group were better than in the non-stimulation group. The results indicate that deacetyl chitin conduit bridging combined with temporary electrical stimulation can promote peripheral nerve repair. PMID:25206762

Zhang, Zhongli; Li, Xin; Zuo, Songjie; Xin, Jie; Zhang, Peixun

2014-01-01

396

Structure-based analysis of domain function of chitin oligosaccharide deacetylase from Vibrio parahaemolyticus.  

PubMed

The X-ray crystal structure of chitin oligosaccharide deacetylase from Vibrio parahaemolyticus (Vp-COD) was determined at an 1.35 Å resolution. The amino acid sequence and structure of Vp-COD show that the enzyme comprises one polysaccharide deacetylase domain (PDD) and two carbohydrate-binding domains (CBDs). On the basis of a chitin-binding assay with Vp-COD and its CBDs-deleted mutant, it was confirmed that CBDs can adhere to chitin. The catalytic activity of the CBDs-deleted mutant was only mildly depressed compared with that of Vp-COD, indicating that CBDs are unlikely to affect the configuration of the active center residues in active site of PDD. PMID:25479092

Hirano, Takako; Sugiyama, Kanako; Sakaki, Yuta; Hakamata, Wataru; Park, Sam-Yong; Nishio, Toshiyuki

2015-01-01

397

Improvement in the quality of hematopoietic prostaglandin D synthase crystals in a microgravity environment  

PubMed Central

Human hematopoietic prostaglandin synthase, one of the better therapeutic target enzymes for allergy and inflammation, was crystallized with 22 inhibitors and in three inhibitor-free conditions in microgravity. Most of the space-grown crystals showed better X-ray diffraction patterns than the terrestrially grown ones, indicating the advantage of a microgravity environment on protein crystallization, especially in the case of this protein. PMID:21169700

Tanaka, Hiroaki; Tsurumura, Toshiharu; Aritake, Kosuke; Furubayashi, Naoki; Takahashi, Sachiko; Yamanaka, Mari; Hirota, Erika; Sano, Satoshi; Sato, Masaru; Kobayashi, Tomoyuki; Tanaka, Tetsuo; Inaka, Koji; Urade, Yoshihiro

2011-01-01

398

Micro-CT Imaging of Denatured Chitin by Silver to Explore Honey Bee and Insect Pathologies  

PubMed Central

Background Chitin and cuticle coatings are important to the environmental and immune defense of honey bees and insect pollinators. Pesticides or environmental effects may target the biochemistry of insect chitin and cuticle coating. Denaturing of chitin involves a combination of deacetylation, intercalation, oxidation, Schweiger-peeling, and the formation of amine hydrochloride salt. The term “denatured chitin” calls attention to structural and property changes to the internal membranes and external carapace of organisms so that some properties affecting biological activities are diminished. Methodology/Principal Findings A case study was performed on honey bees using silver staining and microscopic computer-tomographic x-ray radiography (micro-CT). Silver nitrate formed counter-ion complexes with labile ammonium cations and reacted with amine hydrochloride. Silver was concentrated in the peritrophic membrane, on the abdomen, in the glossa, at intersegmental joints (tarsi), at wing attachments, and in tracheal air sacs. Imaged mono-esters and fatty acids from cuticle coating on external surfaces were apparently reduced by an alcohol pretreatment. Conclusions/Significance The technique provides 3-dimensional and sectional images of individual honey bees consistent with the chemistries of silver reaction and complex formation with denatured chitin. Environmental exposures and influences such as gaseous nitric oxide intercalant, trace oxidants such as ozone gas, oligosachharide salt conversion, exposure to acid rain, and chemical or biochemical denaturing by pesticides may be studied using this technique. Peritrophic membranes, which protect against food abrasion, microorganisms, and permit efficient digestion, were imaged. Apparent surface damage to the corneal lenses of compound eyes by dilute acid exposure consistent with chitin amine hydrochloride formation was imaged. The technique can contribute to existing insect pathology research, and may provide an additional tool for research on CCD. PMID:22110654

Butzloff, Peter R.

2011-01-01