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171.
Mathematical modelling of aliphatic glucosinolate chain length distribution in Arabidopsis thaliana leaves 总被引:1,自引:0,他引:1
Beate Knoke Susanne Textor Jonathan Gershenzon Stefan Schuster 《Phytochemistry Reviews》2009,8(1):39-51
Aliphatic glucosinolates are a major class of defensive secondary metabolites in plants that are mostly derived from methionine.
Occurring in different chain lengths, they show a structural diversity arising from the variable number of chain elongation
cycles taking place during their biosynthesis. The key enzymes in determining glucosinolate chain length are the methylthioalkylmalate
(MAM) synthases, MAM1 and MAM3, with MAM3 showing a broader substrate specificity than MAM1. A comparison of the measurements
of wild type and MAM1 knockout mutant plants shows the following distinct changes in glucosinolate chain length profiles:
MAM3 knockout mutants on the contrary differ from wild type plants by a pronounced abundance of the second shortest glucosinolate
and the depletion of the two longest glucosinolates. To clarify the contribution of the multifunctional enzymes MAM1 and MAM3
to the glucosinolate profile of Arabidopsis thaliana leaves, we simulated glucosinolate biosynthesis in a kinetic model, taking into account the structure of the pathway and
measured enzymatic properties. The predicted glucosinolate profiles show all characteristics of the actual differences between
wild-type and MAM1 mutants or MAM3 mutants, respectively. The model strongly supports experimental indications that the two MAM activities are not independent
of each other. In particular, it showed that an elevated expression of MAM3 in the MAM1 mutant is critical in determining the glucosinolate profile of this plant line. The simulation was critical for this finding
since it allowed us to assess the individual effects of two processes—the knocking out of MAM1 and the overexpression of MAM3—that
are difficult to separate experimentally. 相似文献
(1) | a reversal of the relative proportions of the two shortest glucosinolates, |
(2) | a significant increase in the concentration of the longest glucosinolate, |
(3) | an increase in total glucosinolate content in the mutant. |
172.
Among more than 50 isolates ofBacillus thuringiensis Berliner (B.t.) tested, 7 incited 100% mortality when 2nd instar larvae ofSpodoptera littoralis Boisduval were fed on alfalfa leaves dipped in a spore-crystal suspension of 108 colony forming units/ml. Among those isolates,B.t. 24 demonstrated the highest activity. Larvae of instars 1 and 2 were the most susceptible toB.t. Susceptibility decreased with larval development. However, larvae of all instars were killed by isolateB.t. 24. Larvae that survived after feeding withB.t. 24 were retarded and fed less. Their weight relative to the controls was lower as the spore concentration on the leaves on which they fed was higher. Survival of the spores in the field dropped drastically to 2% after 4 days. Insecticidal activity of the sprayed suspension on those leaves, however, remained significant.B.t. 24 was also effective against larvae on cotton plants in the greenhouse and in a preliminary field experiment. Numbers of colony forming units recovered from leaves dipped in suspension of various spore concentrations showed significant correlation with the initial concentrations as did sprayed leaves. However, colony forming units recovered from sprayed leaves were 5–7.5 fold lower than from dipped leaves. Dipped cotton leaves showed 3.1×10?5 ml attached to 1 mm2 leaf surface whereas sprayed ones had 6×10?6 ml. Those data are important for the determination of spore concentrations in suspensions required for spraying. The isolateB.t. 24 was serotyped byH. de Barjac as H-6B. thuringiensis entomocidus. 相似文献
173.
Cornelia Schuster Helen Brosi Katja Stifter Bernhard O. Boehm Reinhold Schirmbeck 《PloS one》2013,8(8)
Coinhibitory PD-1/PD-L1 (B7-H1) interactions provide critical signals for the regulation of autoreactive T-cell responses. We established mouse models, expressing the costimulator molecule B7.1 (CD80) on pancreatic beta cells (RIP-B7.1 tg mice) or are deficient in coinhibitory PD-L1 or PD-1 molecules (PD-L1−/− and PD-1−/− mice), to study induction of preproinsulin (ppins)-specific CD8 T-cell responses and experimental autoimmune diabetes (EAD) by DNA-based immunization. RIP-B7.1 tg mice allowed us to identify two CD8 T-cell specificities: pCI/ppins DNA exclusively induced Kb/A12–21-specific CD8 T-cells and EAD, whereas pCI/ppinsΔA12–21 DNA (encoding ppins without the COOH-terminal A12–21 epitope) elicited Kb/B22–29-specific CD8 T-cells and EAD. Specific expression/processing of mutant ppinsΔA12–21 (but not ppins) in non-beta cells, targeted by intramuscular DNA-injection, thus facilitated induction of Kb/B22–29-specific CD8 T-cells. The A12–21 epitope binds Kb molecules with a very low avidity as compared with B22–29. Interestingly, immunization of coinhibition-deficient PD-L1−/− or PD-1−/− mice with pCI/ppins induced Kb/A12–21-monospecific CD8 T-cells and EAD but injections with pCI/ppinsΔA12–21 did neither recruit Kb/B22–29-specific CD8 T-cells into the pancreatic target tissue nor induce EAD. PpinsΔA12–21/(Kb/B22–29)-mediated EAD was efficiently restored in RIP-B7.1+/PD-L1−/− mice, differing from PD-L1−/− mice only in the tg B7.1 expression in beta cells. Alternatively, an ongoing beta cell destruction and tissue inflammation, initiated by ppins/(Kb/A12–21)-specific CD8 T-cells in pCI/ppins+pCI/ppinsΔA12–21 co-immunized PD-L1−/− mice, facilitated the expansion of ppinsΔA12–21/(Kb/B22–29)-specific CD8 T-cells. CD8 T-cells specific for the high-affinity Kb/B22–29- (but not the low-affinity Kb/A12–21)-epitope thus require stimulatory ´help from beta cells or inflamed islets to expand in PD-L1-deficient mice. The new PD-1/PD-L1 diabetes models may be valuable tools to study under well controlled experimental conditions distinct hierarchies of autoreactive CD8 T-cell responses, which trigger the initial steps of beta cell destruction or emerge during the pathogenic progression of EAD. 相似文献
174.
175.
Chloroplast PNPase exists as a homo-multimer enzyme complex that is distinct from the Escherichia coli degradosome. 下载免费PDF全文
S Baginsky A Shteiman-Kotler V Liveanu S Yehudai-Resheff M Bellaoui R E Settlage J Shabanowitz D F Hunt G Schuster W Gruissem 《RNA (New York, N.Y.)》2001,7(10):1464-1475
In Escherichia coli, the exoribonuclease polynucleotide phosphorylase (PNPase), the endoribonuclease RNase E, a DEAD-RNA helicase and the glycolytic enzyme enolase are associated with a high molecular weight complex, the degradosome. This complex has an important role in processing and degradation of RNA. Chloroplasts contain an exoribonuclease homologous to E. coli PNPase. Size exclusion chromatography revealed that chloroplast PNPase elutes as a 580-600 kDa complex, suggesting that it can form an enzyme complex similar to the E. coli degradosome. Biochemical and mass-spectrometric analysis showed, however, that PNPase is the only protein associated with the 580-600 kDa complex. Similarly, a purified recombinant chloroplast PNPase also eluted as a 580-600 kDa complex after gel filtration chromatography. These results suggest that chloroplast PNPase exists as a homo-multimer complex. No other chloroplast proteins were found to associate with chloroplast PNPase during affinity chromatography. Database analysis of proteins homologous to E. coli RNase E revealed that chloroplast and cyanobacterial proteins lack the C-terminal domain of the E. coli protein that is involved in assembly of the degradosome. Together, our results suggest that PNPase does not form a degradosome-like complex in the chloroplast. Thus, RNA processing and degradation in this organelle differ in several respects from those in E. coli. 相似文献
176.
Thompson JE Kutateladze TG Schuster MC Venegas FD Messmore JM Raines RT 《Bioorganic chemistry》1995,23(4):471-481
Bovine pancreatic ribonuclease A (RNase A) catalyzes the cleavage of the P-O(5') bond in RNA. Although this enzyme has been the object of much landmark work in bioorganic chemistry, the nature of its rate-limiting transition state and its catalytic rate enhancement had been unknown. Here, the value of k(cat)/K(m) for the cleavage of UpA by wild-type RNase A was found to be inversely related to the concentration of added glycerol. In contrast, the values of k(cat)/K(m) for the cleavage of UpA by a sluggish mutant of RNase A and the cleavage of the poor substrate UpOC(6)H(4)-p-NO(2) by wild-type RNase A were found to be independent of glycerol concentration. Yet, UpA cleavage by the wild-type and mutant enzymes was found to have the same dependence on sucrose concentration, indicating that catalysis of UpA cleavage by RNase A is limited by desolvation. The rate of UpA cleavage by RNase A is maximal at pH 6.0, where k(cat) = 1.4 × 10(3) s(-1) and k(cat)/K(m) = 2.3 × 10(6) M(-1)s(-1) at 25°C. At pH 6.0 and 25°C, the uncatalyzed rate of [5,6-(3)H]Up[3,5,8-(3)H]A cleavage was found to be k(uncat) = 5 × 10(-9) s(-1) (t(1/2) = 4 years). Thus, RNase A enhances the rate of UpA cleavage by 3 × 10(11)-fold by binding to the transition state for P-O(5') bond cleavage with a dissociation constant of <2 × 10(-15) M. 相似文献
177.
Root-knot nematodes, Meloidogyne incognita, induced lumps of callus tissue on the cambial surfaces of peeled tobacco stem segments cultured in vitro. Except for a layer 1 to 3 cells thick, callus was limited to the basal ends of control segments. Indole-3-acetic acid (IAA) applied in agar blocks to the centers of stem segments, when it had any effect on the cambial surface, induced streaks of callus extending from the blocks toward the basal ends of the segments. IAA in agar blocks also increased callus growth at the basal ends of the segments, increased the growth of pith on the undersides of the segments, promoted root initiation, but inhibited bud initiation. Nematodes produced none of these effects, nor did they change the type of organs induced by various concentrations of IAA in the medium. Callus tissue did grow on the cambial surface of stem segments surrounding agar blocks containing 2,3,5-triiodobenzoic acid, an inhibitor of polar auxin transport. Paraffin sections showed that the nematodes were confined to the callus tissue on the cambial surfaces of the segments. Except for occasional syncytia and areas of cell division, nematode-induced callus was composed of thin-walled, irregularly shaped cells arising from the cambium. Differences between the responses of tobacco stem segments to root-knot nematodes and IAA-agar blocks indicate that auxins were not freed from the plant tissue nor secreted by the nematodes. Instead, it is suggested that nematodes enabled the tissue to retain and use endogenous auxins that otherwise would have been transported to the basal ends of the segments. 相似文献
178.
Biophysical characterization of a designed TMV coat protein mutant, R46G, that elicits a moderate hypersensitivity response in Nicotiana sylvestris 下载免费PDF全文
Toedt JM Braswell EH Schuster TM Yphantis DA Taraporewala ZF Culver JN 《Protein science : a publication of the Protein Society》1999,8(2):261-270
The hypersensitivity resistance response directed by the N' gene in Nicotiana sylvestris is elicited by the tobacco mosaic virus (TMV) coat protein R46G, but not by the U1 wild-type TMV coat protein. In this study, the structural and hydrodynamic properties of R46G and wild-type coat proteins were compared for variations that may explain N' gene elicitation. Circular dichroism spectroscopy reveals no significant secondary or tertiary structural differences between the elicitor and nonelicitor coat proteins. Analytical ultracentrifugation studies, however, do show different concentration dependencies of the weight average sedimentation coefficients at 4 degrees C. Viral reconstitution kinetics at 20 degrees C were used to determine viral assembly rates and as an initial assay of the rate of 20S formation, the obligate species for viral reconstitution. These kinetic results reveal a decreased lag time for reconstitution performed with R46G that initially lack the 20S aggregate. However, experiments performed with 20S initially present reveal no detectable differences indicating that the mechanism of viral assembly is similar for the two coat protein species. Therefore, an increased rate of 20S formation from R46G subunits may explain the differences in the viral reconstitution lag times. The inferred increase in the rate of 20S formation is verified by direct measurement of the 20S boundary as a function of time at 20 degrees C using velocity sedimentation analysis. These results are consistent with the interpretation that there may be an altered size distribution and/or lifetime of the small coat protein aggregates in elicitors that allows N. sylvestris to recognize the invading virus. 相似文献
179.
T. L. Hamilton R. J. Bovee V. Thiel S. R. Sattin W. Mohr I. Schaperdoth K. Vogl W. P. Gilhooly III T. W. Lyons L. P. Tomsho S. C. Schuster J. Overmann D. A. Bryant A. Pearson J. L. Macalady 《Geobiology》2014,12(5):451-468
Mahoney Lake represents an extreme meromictic model system and is a valuable site for examining the organisms and processes that sustain photic zone euxinia (PZE). A single population of purple sulfur bacteria (PSB) living in a dense phototrophic plate in the chemocline is responsible for most of the primary production in Mahoney Lake. Here, we present metagenomic data from this phototrophic plate – including the genome of the major PSB, as obtained from both a highly enriched culture and from the metagenomic data – as well as evidence for multiple other taxa that contribute to the oxidative sulfur cycle and to sulfate reduction. The planktonic PSB is a member of the Chromatiaceae, here renamed Thiohalocapsa sp. strain ML1. It produces the carotenoid okenone, yet its closest relatives are benthic PSB isolates, a finding that may complicate the use of okenone (okenane) as a biomarker for ancient PZE. Favorable thermodynamics for non‐phototrophic sulfide oxidation and sulfate reduction reactions also occur in the plate, and a suite of organisms capable of oxidizing and reducing sulfur is apparent in the metagenome. Fluctuating supplies of both reduced carbon and reduced sulfur to the chemocline may partly account for the diversity of both autotrophic and heterotrophic species. Collectively, the data demonstrate the physiological potential for maintaining complex sulfur and carbon cycles in an anoxic water column, driven by the input of exogenous organic matter. This is consistent with suggestions that high levels of oxygenic primary production maintain episodes of PZE in Earth's history and that such communities should support a diversity of sulfur cycle reactions. 相似文献
180.
Schuster D Kowalik D Kirchmair J Laggner C Markt P Aebischer-Gumy C Ströhle F Möller G Wolber G Wilckens T Langer T Odermatt A Adamski J 《The Journal of steroid biochemistry and molecular biology》2011,125(1-2):148-161
17β-Hydroxysteroid dehydrogenase type 3 and 5 (17β-HSD3 and 17β-HSD5) catalyze testosterone biosynthesis and thereby constitute therapeutic targets for androgen-related diseases or endocrine-disrupting chemicals. As a fast and efficient tool to identify potential ligands for 17βHSD3/5, ligand- and structure-based pharmacophore models for both enzymes were developed. The models were evaluated first by in silico screening of commercial compound databases and further experimentally validated by enzymatic efficacy tests of selected virtual hits. Among the 35 tested compounds, 11 novel inhibitors with distinct chemical scaffolds, e.g. sulfonamides and triazoles, and with different selectivity properties were discovered. Thereby, we provide several potential starting points for further 17β-HSD3 and 17β-HSD5 inhibitor development. Article from the Special issue on Targeted Inhibitors. 相似文献