首页 | 本学科首页   官方微博 | 高级检索  
相似文献
 共查询到20条相似文献,搜索用时 15 毫秒
1.
The motile zoospores of the damping-off pathogen Aphanomyces cochlioides aggregate on host plants (e.g., sugar beet, spinach) guided by the host-specific plant signal cochliophilin A before infection. To assess the potential role of secondary metabolites in nonhost resistance, acetone extracts of 200 nonhost traditional medicinal plants from Chinese and Bangladeshi origins were tested for the motility behaviour of A. cochlioides zoospores using a particle bioassay method. Nearly one third of the tested plant extracts exhibited diverse deleterious activities such as repellent, stimulant, motility halting and lysis against A. cochlioides zoospores. Among these active plants, an extract of the Chinese medicinal plant Dalbergia odorifera displayed potent repellent activity toward zoospores. Chromatographic separation of D. odorifera constituents revealed that the repellent activity was regulated by the cumulative effect of three motility-affecting isoflavonoids, viz. (+/-)-medicarpin (repellent at 150 microg/ml), (-)-claussequinone (stimulant at 100 microg/ml) and formononetin (stimulant and attractant at 50 microg/ml). A mixture (1:1:1, w/w/w) of these three compounds exhibited only repellent activity toward zoospores at a concentration lower than 50 microg/ml. These results suggest that nonhost plants might possess potential bioactive secondary metabolites to ward off zoosporic phytopathogens.  相似文献   

2.
The EtOAc soluble constituents of the unripe fruits of Ginkgo biloba showed motility inhibition followed by lysis of zoospores of the phytopathogenic Aphanomyces cochlioides. We purified 22:1-omega7-anacardic acid (1), 24:1-omega9-anacardic acid (2) and 22:0-anacardic acid (3), together with other related compounds, 21:1-omega7-cardol (4) and 21:1-omega7-cardanol (5) from the crude extracts of Ginkgo fruits. Amongst them, compound 1 was a major active agent in quality and quantity, and showed potent motility inhibition (98% in 30 min) followed by lysis (55% in 3 h) of the zoospores at 1 x 10(-7) M. The 2-O-methyl derivative (1-c) of 1 displayed antibacterial activity against Bacillus subtilis, but practically inactive to Escherichia coli. A brief study on structure-activity relationships revealed that a carboxyl group on the aromatic ring and an unsaturated side chain in the anacardic acid derivative are important for strong motility inhibitory and lytic activities against the zoospore.  相似文献   

3.
Cochliophilin A (5-hydroxy-6,7-methylenedioxyflavone), a root releasing host-specific plant signal triggers chemotaxis and subsequent morphological changes in pathogenic Aphanomyces cochlioides zoospores before host penetration. The present study illustrates time-course changing patterns of cytoskeletal filamentous actin (F-actin) organization in the zoospores of A. cochlioides during rapid morphological changes (encystment and germination) after exposure to cochliophilin A. Confocal laser scanning microscopic analysis revealed that F-actin microfilaments remained concentrated at ventral groove and diffusely distributed in peripheral cytoplasm of the zoospore. These microfilaments dramatically rearranged and changed into granular F-actin plaques interconnected with fine arrays during encystment. A large patch of actin arrays accumulated at one pole of the cystospores just before germination. Then the actin plaques moved to the emerging germ tube where a distinct cap of microfilaments was seen at the tip of the emerging hypha. Zoospores treated with an inhibitor of F-actin polymerization, latrunculin B or motility halting and regeneration inducing compound nicotinamide, displayed different patterns of F-actin in both zoospores and cystospores than those obtained by the induction of cochliophilin A. Collectively, these results indicate that the host-specific plant signal cochliophilin A triggers a dynamic polymerization/depolymerization of F-actin in pathogenic A. cochlioides zoospores during early events of plant-peronosporomycete interactions.  相似文献   

4.
Screening chemical compounds, we found that a xenoestrogen, bisphenol A, showed potent repellent activity against the zoospores of Aphanomyces cochlioides. Based on this finding, we tested a number of androgenic and estrogenic compounds (e.g. testosterone, progesterone, estradiols, diethylstilbestrol, estrone, estriol, pregnenolone, dienestrol etc.) on the motility behavior of A. cochlioides zoospores. Interestingly, most of the estrogenic compounds exhibited potent repellent activity (1 microg/ml or less by the "particle method") toward the motile zoospores of A. cochlioides. We derivatized some of the estrogens and discussed the relationship between the structure of active molecules and their repellent activity. Apparently, aromatization of the A ring with a free hydroxyl group at C-3 position of a steroidal structure is necessary for higher repellent activity. Interestingly, methylation of diethylstilbestrol (DES) yielded completely different activity i.e. both mono- and di-methyl ethers of DES showed attractant activity. Moreover, the attracted zoospores were encysted and then germinated in the presence of di-methyl ether of DES. The potential usefulness of this repellent test is discussed for the detection of estrogenic activity of naturally occurring compounds, and the possible role of phytoestrogens in host/parasite interactions. So far, this will be the first report of repellent activity of estrogenic compounds toward trivial fungal zoospores.  相似文献   

5.
We previously demonstrated that xanthobaccin A from the rhizoplane bacterium Lysobacter sp. strain SB-K88 suppresses damping-off disease caused by Pythium sp. in sugar beet. In this study we focused on modes of Lysobacter sp. strain SB-K88 root colonization and antibiosis of the bacterium against Aphanomyces cochlioides, a pathogen of damping-off disease. Scanning electron microscopic analysis of 2-week-old sugar beet seedlings from seeds previously inoculated with SB-K88 revealed dense colonization on the root surfaces and a characteristic perpendicular pattern of Lysobacter colonization possibly generated via development of polar, brush-like fimbriae. In colonized regions a semitransparent film apparently enveloping the root and microcolonies were observed on the root surface. This Lysobacter strain also efficiently colonized the roots of several plants, including spinach, tomato, Arabidopsis thaliana, and Amaranthus gangeticus. Plants grown from both sugar beet and spinach seeds that were previously treated with Lysobacter sp. strain SB-K88 displayed significant resistance to the damping-off disease triggered by A. cochlioides. Interestingly, zoospores of A. cochlioides became immotile within 1 min after exposure to a SB-K88 cell suspension, a cell-free supernatant of SB-K88, or pure xanthobaccin A (MIC, 0.01 microg/ml). In all cases, lysis followed within 30 min in the presence of the inhibiting factor(s). Our data indicate that Lysobacter sp. strain SB-K88 has a direct inhibitory effect on A. cochlioides, suppressing damping-off disease. Furthermore, this inhibitory effect of Lysobacter sp. strain SB-K88 is likely due to a combination of antibiosis and characteristic biofilm formation at the rhizoplane of the host plant.  相似文献   

6.
Peng Y  Sartini D  Pozzi V  Wilk D  Emanuelli M  Yee VC 《Biochemistry》2011,50(36):7800-7808
Nicotinamide N-methyltransferase (NNMT) catalyzes the N-methylation of nicotinamide, pyridines, and other analogues using S-adenosyl-l-methionine as donor. NNMT plays a significant role in the regulation of metabolic pathways and is expressed at markedly high levels in several kinds of cancers, presenting it as a potential molecular target for cancer therapy. We have determined the crystal structure of human NNMT as a ternary complex bound to both the demethylated donor S-adenosyl-l-homocysteine and the acceptor substrate nicotinamide, to 2.7 ? resolution. These studies reveal the structural basis for nicotinamide binding and highlight several residues in the active site which may play roles in nicotinamide recognition and NNMT catalysis. The functional importance of these residues was probed by mutagenesis. Of three residues near the nicotinamide's amide group, substitution of S201 and S213 had no effect on enzyme activity while replacement of D197 dramatically decreased activity. Substitutions of Y20, whose side chain hydroxyl interacts with both the nicotinamide aromatic ring and AdoHcy carboxylate, also compromised activity. Enzyme kinetics analysis revealed k(cat)/K(m) decreases of 2-3 orders of magnitude for the D197A and Y20A mutants, confirming the functional importance of these active site residues. The mutants exhibited substantially increased K(m) for both NCA and AdoMet and modestly decreased k(cat). MD simulations revealed long-range conformational effects which provide an explanation for the large increase in K(m)(AdoMet) for the D197A mutant, which interacts directly only with nicotinamide in the ternary complex crystal structure.  相似文献   

7.
Aphanomyces cochlioides zoospores show chemotaxis to cochliophilin A (5-hydroxy-6,7-methylenedioxyflavone, 1), a host derived attractant, and also respond to 5,7-dihydroxyflavone (2) known as an equivalent chemoattractant. To investigate the chemotactic receptors in the zoospores, we designed photoaffinity probes 4'-azido-5,7-dihydroxyflavone (3) and 4'-azido-7-O-biotinyl-5-hydroxyflavone (4) considering chemical structure of 2. Both 3 and 4 had zoospore attractant activity which was competitive with that of 1. When zoospores were treated with the biotinylated photoaffinity probe followed by UV irradiation and streptavidin-gold or peroxidase-conjugated streptavidin, probe-labeled proteins were detected on the cell membrane. This result indicated that the 1-specific-binding proteins, a candidate for hypothetical cochliophilin A receptor, were localized on the cell membrane of the zoospores. This is the first experimental evidence of flavonoid-binding proteins being present in zoospores, using chemically synthesized azidoflavone as photoaffinity-labeling reagent.  相似文献   

8.
A bioassay-guided survey of spinach leaf constituents resulted in 5,4'-dihydroxy-3,3'-dimethoxy-6,7-methylenedioxyflavone being identified as the third naturally-occurring attractant in the host plant toward the zoospores of its pathogen, Aphanomyces cochlioides. The isolate showed attracting activity around Chromosorb W AW particles (60-80 mesh) coated with a 10(-5) M solution in a zoospore suspension. However, this activity was 1/100-1/1000 less than that of cochliophilin A, an attractant in the roots of spinach. Bioassays with the present isolate and related compounds revealed that 5,3',4'-trihydroxy-3-methoxy-6,7-methylenedioxyflavone did not possess attractant activity, but rather weak antagonistic activity toward the former two attractants from spinach.  相似文献   

9.
Neutrophils constitute the first line of host defense against invading microorganisms. Yet their removal from the inflammatory environment is fundamental for injury restraint and resolution of inflammation. Nicotinamide, a component of vitamin B(3), is known to modulate cell survival. In this study, we assessed the influence of nicotinamide on neutrophil apoptosis, both in vitro and in vivo in a mouse model of endotoxin-induced lung inflammation. In vitro, nicotinamide promoted apoptosis of human blood neutrophils in a dose-dependent manner in the presence of the apoptosis inhibitors granulocyte colony-stimulating factor and granulocyte/macrophage colony-stimulating factor. The highest concentration of nicotinamide completely neutralized the pro-survival effect of granulocyte (macrophage) colony-stimulating factor. Nicotinamide proapoptotic effect was associated with enhanced caspase-3 activity. In addition, nicotinamide slightly reduced neutrophil chemotaxis in vitro. In vivo, pulmonary nicotinamide delivery decreased the levels of cellular and biochemical inflammation markers and increased the percentage of apoptotic neutrophils in bronchoalveolar lavages. Our findings suggest that nicotinamide is an apoptotic stimulus for neutrophils, thereby contributing to the resolution of neutrophilic inflammation in the lungs.  相似文献   

10.
Nicotinamide phosphoribosyltransferase, (EC 2.4.2.12) was examined in extracts of diploid human fibroblasts grown in culture. The enzyme was found to have an apparent Km for nicotinamide of 1.6 × 10?6M, to be specific for nicotinamide, stimulated by adenosine triphosphate (ATP) and inhibited by nicotinamide adenine dinucleotide (NAD). In these respects it is very similar to rat liver nicotinamide phosphoribosyltransferase but not like the enzyme previously observed in human tissue extracts which had a Km for nicotinamide of approximately 0.1 M and was insensitive to ATP. Discovery of this enzyme activity supports previous studies using radiolabeled nicotinamide which show that human fibroblasts can incorporate nicotinamide into NAD directly through nicotinamide mononucleotide.  相似文献   

11.
Three 4-thiazolidinones, two with nicotinamide (NAT1 and NAT2) and one with 4-chlorophenoxyacetamide (PAT1) side chains were evaluated for their hypolipidaemic, hypoglycaemic activity in Swiss albino mice fed a high-fat diet along with fructose administered in drinking water. NAT1 and PAT caused reduction of elevated triglycerides, cholesterol and glucose; NAT2 was effective only against triglycerides. Nicotinamide side chain might have contributed to the lipid lowering effect of both NAT1 and NAT2, but the bulky group of the latter could have affected proper binding to the receptor sites, making it ineffective against elevated cholesterol. On the other hand, the 4-chlorophenoxyacetamide side chain of PAT might have exerted powerful hypolipidaemic activity, despite the bulky substitution at C2. As antioxidants, NAT2 and PAT1 showed superior activity, compared to NAT1. The thiazolidinone ring might be responsible for the lipid lowering effect, which is however, modified by the type of substitutions at C2 and N of the ring. Detailed study is warranted to explain the mechanism of action of these compounds as also to make more potent ones.  相似文献   

12.
Utilization and metabolism of NAD by Haemophilus parainfluenzae   总被引:2,自引:0,他引:2  
The utilization of exogenous nicotinamide adenine dinucleotide (NAD) by Haemophilus parainfluenzae was studied in suspensions of whole cells using radiolabelled NAD, nicotinamide mononucleotide (NMN), and nicotinamide ribonucleoside (NR). The utilization of these compounds by H. parainfluenzae has the following characteristics. (1) NAD is not taken up intact, but rather is degraded to NMN or NR prior to internalization. (2) Uptake is carrier-mediated and energy-dependent with saturation kinetics. (3) There is specificity for the beta-configuration of the glycopyridine linkage. (4) An intact carboxamide groups is required on the pyridine ring. The intracellular metabolism of NAD was studied in crude cell extracts and in whole cells using carbonyl-14C-labelled NR, NMN, NAD, nicotinamide, and nicotinic acid as substrates in separate experiments. A synthetic pathway from NR through NMN to NAD that requires Mg2+ and ATP was demonstrated. Nicotinamide was found as an end-product of NAD degradation. Nicotinic acid mononucleotide and nicotinic acid adenine dinucleotide were not found as intermediates. The NAD synthetic pathway in H. parainfluenzae differs from the Preiss-Handler pathway and the pyridine nucleotide cycles described in other bacteria.  相似文献   

13.
Nicotinamide was shown to inhibit deoxyglucose uptake in three diverse differentiated cell lines. In 3T3-L1 fat cells, nicotinamide equally inhibited basal and insulin stimulated deoxyglucose uptake. Inhibition by nicotinamide was non-competitive. A variety of inhibitors of ADP-ribosylation blocked deoxyglucose uptake while some analogs with no activity against ADP-ribose synthetase also had little effect on deoxyglucose uptake. These findings should be taken into account when inhibitors of ADP-ribosylation are used with intact cells.  相似文献   

14.
Nicotinamide adenine dinucleotide synthetase (NadE) is an essential enzyme for bacterial pathogens and is thus a promising antibacterial target. It catalyzes the conversion of nicotinic acid adenine dinucleotide to nicotinamide adenine dinucleotide. Changes in chemical shifts that occur in the nicotinic acid ring as it is converted to nicotinamide can be used for monitoring the reaction. A robust nuclear magnetic resonance-based activity assay was developed using robotically controlled reaction initiation and quenching. The single-enzyme assay has less potential for false positives compared to a coupled activity assay and is especially well suited to the high concentration of compounds in fragment screens. The assay has been used to screen fragment libraries for NadE inhibitors.  相似文献   

15.
Readily synthesized nicotinamide adenine dinucleotide (NAD(+)) analogues have been used to investigate aspects of the cyclization of NAD(+) to cyclic adenosine 5'-O-diphosphate ribose (cADPR) catalyzed by the enzyme adenosine 5'-O-diphosphate (ADP) ribosyl cyclase and to produce the first potent inhibitors of this enzyme. In all cases, inhibition of Aplysia californica cyclase by various substrate analogues was found to be competitive while inhibition by nicotinamide exhibited mixed-behavior characteristics. Nicotinamide hypoxanthine dinucleotide (NHD(+)), nicotinamide guanine dinucleotide (NGD(+)), C1'-m-benzamide adenine dinucleotide (Bp(2)A), and C1'-m-benzamide nicotinamide dinucleotide (Bp(2)N) were found to be nanomolar potency inhibitors with inhibition constants of 70, 143, 189, and 201 nM, respectively. However, NHD(+) and NGD(+) are also known substrates and are slowly converted to cyclic products, thus preventing their further use as inhibitors. The symmetrical bis-nucleotides, bis-adenine dinucleotide (Ap(2)A), bis-hypoxanthine dinucleotide (Hp(2)H), and bis-nicotinamide dinucleotide (Np(2)N), exhibited micromolar competitive inhibition, with Ap(2)A displaying the greatest affinity for the enzyme. 2',3'-Di-O-acetyl nicotinamide adenine dinucleotide (AcONAD(+)) was not a substrate for the A. californica cyclase but also displayed some inhibition at a micromolar level. Finally, inhibition of the cyclase by adenosine 5'-O-diphosphate ribose (ADPR) and inosine 5'-O-diphosphate ribose (IDPR) was observed at millimolar concentration. The nicotinamide aromatic ring appears to be the optimal motif required for enzymatic recognition, while modifications of the 2'- and 3'-hydroxyls of the nicotinamide ribose seem to hamper binding to the enzyme. Stabilizing enzyme/inhibitor interactions and the inability of the enzyme to release unprocessed material are both considered to explain nanomolar inhibition. Recognition of inhibitors by other ADP ribosyl cyclases has also been investigated, and this study now provides the first potent nonhydrolyzable sea urchin ADP ribosyl cyclase and cADPR hydrolase inhibitor Bp(2)A, with inhibition observed at the micromolar and nanomolar level, respectively. The benzamide derivatives did not inhibit CD38 cyclase or hydrolase activity when NGD(+) was used as substrate. These results emphasize the difference between CD38 and other enzymes in which the cADPR cyclase activity predominates.  相似文献   

16.
Nicotinamide inhibited both germinal vesicle breakdown (GVBD) and polar body formation (PBF) in surf clam and starfish oocytes. In the surf clam nicotinamide at 0.3 mM completely blocked PBF in the fertilized oocytes. For blockage of GVBD higher concentration was required. In the starfish, nicotinamide (30 mM) prevented PBF but not GVBD, when added 7 min after the commencement of 1-methyladenine (1-MeAde) administration. These results suggest that PBF is blocked by nicotinamide independent of its effect on GVBD. In the case of starfish, NAD+was more effective than nicotinamide in inhibiting oocyte maturation. Nicotinamide also blocked GVBD induced by microinjection of the cytoplasm containing maturation-promoting factor (MPF) obtained from 1-MeAde-treatcd oocytes. These results suggest that nicotinamide prevents the action of MPF rather than inhibiting the interaction of 1-McAde with cell membrane or the induction of MPF.  相似文献   

17.
Cochliophilin A (5-hydroxy-6,7-methylenedioxyflavone, 1), known as a host-specific attractant towards the zoospores of Aphanomyces cochlioides, a cause of root rot and damping-off diseases of Chenopodiaceae, was found in the Amaranthaceae plant, Celosia cristata, that is susceptible to the pathogen. The content of 1 in Celosia seedlings was quantified as 1.4 microg/g fresh weight. A new isoflavone, cristatein (5-hydroxy-6-hydroxymethyl-7,2'-dimethoxyisoflavone, 2), and five known flavonoids were also identified.  相似文献   

18.
NAD(+) is an essential coenzyme for hydride transfer enzymes and a substrate of sirtuins and other NAD(+)-consuming enzymes. Nicotinamide riboside is a recently discovered eukaryotic NAD(+) precursor converted to NAD(+) via the nicotinamide riboside kinase pathway and by nucleosidase activity and nicotinamide salvage. Nicotinamide riboside supplementation of yeast extends replicative life span on high glucose medium. The molecular basis for nicotinamide riboside uptake was unknown in any eukaryote. Here, we show that deletion of a single gene, YOR071C, abrogates nicotinamide riboside uptake without altering nicotinic acid or nicotinamide import. The gene, which is negatively regulated by Sum1, Hst1, and Rfm1, fully restores nicotinamide riboside import and utilization when resupplied to mutant yeast cells. The encoded polypeptide, Nrt1, is a predicted deca-spanning membrane protein related to the thiamine transporter, which functions as a pH-dependent facilitator with a K(m) for nicotinamide riboside of 22 microm. Nrt1-related molecules are conserved in particular fungi, suggesting a similar basis for nicotinamide riboside uptake.  相似文献   

19.
Chloroplast ferredoxin-NADP(+) reductase has a 32,000-fold preference for NADPH over NADH, consistent with its main physiological role of NADP(+) photoreduction for de novo carbohydrate biosynthesis. Although it is distant from the 2'-phosphoryl group of NADP(+), replacement of the C-terminal tyrosine (Tyr(308) in the pea enzyme) by Trp, Phe, Gly, and Ser produced enzyme forms in which the preference for NADPH over NADH was decreased about 2-, 10-, 300-, and 400-fold, respectively. Remarkably, in the case of the Y308S mutant, the k(cat) value for the NADH-dependent activity approached that of the NADPH-dependent activity of the wild-type enzyme. Furthermore, difference spectra of the NAD(+) complexes revealed that the nicotinamide ring of NAD(+) binds at nearly full occupancy in the active site of both the Y308G and Y308S mutants. These results correlate well with the k(cat) values obtained with these mutants in the NADH-ferricyanide reaction. The data presented support the hypothesis that specific recognition of the 2'-phosphate group of NADP(H) is required but not sufficient to ensure a high degree of discrimination against NAD(H) in ferredoxin-NADP(+) reductase. Thus, the C-terminal tyrosine enhances the specificity of the reductase for NADP(H) by destabilizing the interaction of a moiety common to both coenzymes, i.e. the nicotinamide.  相似文献   

20.
NADPH-cytochrome P450 oxidoreductase catalyzes transfer of electrons from NADPH, via two flavin cofactors, to various cytochrome P450s. The crystal structure of the rat reductase complexed with NADP(+) has revealed that nicotinamide access to FAD is blocked by an aromatic residue (Trp-677), which stacks against the re-face of the isoalloxazine ring of the flavin. To investigate the nature of interactions between the nicotinamide, FAD, and Trp-677 during the catalytic cycle, three mutant proteins were studied by crystallography. The first mutant, W677X, has the last two C-terminal residues, Trp-677 and Ser-678, removed; the second mutant, W677G, retains the C-terminal serine residue. The third mutant has the following three catalytic residues substituted: S457A, C630A, and D675N. In the W677X and W677G structures, the nicotinamide moiety of NADP(+) lies against the FAD isoalloxazine ring with a tilt of approximately 30 degrees between the planes of the two rings. These results, together with the S457A/C630A/D675N structure, allow us to propose a mechanism for hydride transfer regulated by changes in hydrogen bonding and pi-pi interactions between the isoalloxazine ring and either the nicotinamide ring or Trp-677 indole ring. Superimposition of the mutant and wild-type structures shows significant mobility between the two flavin domains of the enzyme. This, together with the high degree of disorder observed in the FMN domain of all three mutant structures, suggests that conformational changes occur during catalysis.  相似文献   

设为首页 | 免责声明 | 关于勤云 | 加入收藏

Copyright©北京勤云科技发展有限公司  京ICP备09084417号