scholarly journals Triadimefon, a fungicidal triazole-type P450 inhibitor, induces brassinosteroid deficiency-like phenotypes in plants and binds to DWF4 protein in the brassinosteroid biosynthesis pathway

2003 ◽  
Vol 369 (1) ◽  
pp. 71-76 ◽  
Author(s):  
Tadao ASAMI ◽  
Masaharu MIZUTANI ◽  
Yukihisa SHIMADA ◽  
Hideki GODA ◽  
Nobutaka KITAHATA ◽  
...  
Keyword(s):  
Binding Assay ◽  
Feedback Regulation ◽  
Biosynthesis Pathway ◽  
In Planta ◽  
Biosynthesis Inhibitors ◽  
Target Sites ◽  
Rescue Experiment ◽  
Arabidopsis Cells ◽  
Ga Biosynthesis ◽  
Good Agreement

Triadimefon (Bayleton®), a widely used triazole-type fungicide, affects gibberellin (GA) biosynthesis and 14α-demethylase in sterol biosynthesis. The present study revealed that the phenotype of Arabidopsis treated with triadimefon resembled that of a brassinosteroid (BR)-biosynthesis mutant, and that the phenotype was rescued by brassinolide (BL), the most active BR, partly rescued by GA, and fully rescued by the co-application of BL and GA, suggesting that triadimefon affects both BR and GA biosynthesis. The target sites of triadimefon were investigated using a rescue experiment, feeding triadimefon-treated Arabidopsis BR-biosynthesis intermediates, and a binding assay to expressed DWF4 protein, which is reported to be involved in the BR-biosynthesis pathway. The binding assay indicated that the dissociation constant for triadimefon was in good agreement with the activity in an in planta assay. In the triadimefon-treated Arabidopsis cells, the CPD gene in the BR-biosynthesis pathway was up-regulated, probably due to feedback regulation caused by BR deficiency. These results strongly suggest that triadimefon inhibits the reaction catalysed by DWF4 protein and induces BR deficiency in plants. As triadimefon treatment has proved to be beneficial to plants, this result suggests that BR-biosynthesis inhibitors can be applied to crops.

Daminozide Enhances Vigor and Steviol Glycoside Yield of Stevia (Stevia rebaudiana Bert.) Propagated in the Temporary Immersion Bioreactor

2022 ◽  
Author(s):  
Rizka Tamania Saptari ◽  
Rizkita Rachmi Esyanti ◽  
Riza Arief Putranto
Keyword(s):  
Stevia Rebaudiana ◽  
Feedback Regulation ◽  
Optimum Concentration ◽  
Temporary Immersion Bioreactor ◽  
Temporary Immersion ◽  
Steviol Glycoside ◽  
Kaurenoic Acid ◽  
Biosynthesis Gene ◽  
Inhibitor Treatment ◽  
Ga Biosynthesis

Abstract Stevia (Stevia rebaudiana Bertoni) contains sweet compound widely used as natural sweetener, steviol glycoside (SG). SG is a diterpenoid secondary metabolite synthesized from ent-kaurenoic acid, the same precursor of Gibberellin (GA). Therefore, in this study, a GA inhibitor, Daminozide (0, 10, 20 ppm) was used to block ent-kaurenoic acid conversion towards GA synthesis in attempt to increase SG content of stevia propagated in Temporary Immersion Bioreactor (TIB). Daminozide in 10 mg/L was observed to be the optimum concentration which increased biomass weight and SG content (stevioside and rebaudioside A) up to 40%. The treatment also increased transcripts accumulation of genes enrolled in SG biosynthesis, such as SrKA13H, SrUGT85C2, and SrUGT76G1, indicating SG pathway become more active due to the inhibition of GA pathway. Furthermore, the inhibition of GA was also indicated by the upregulated expression of GA biosynthesis gene (GA3ox) as the result of feedback regulation, and the downregulated expression of GA catabolism gene (GA2ox2) as the result of feed-forward regulation caused by inhibitor treatment.

scholarly journals Allosteric regulation of menaquinone (vitamin K2) biosynthesis in the human pathogen Mycobacterium tuberculosis

2020 ◽  
Vol 295 (12) ◽  
pp. 3759-3770 ◽  
Author(s):  
Ghader Bashiri ◽  
Laura V. Nigon ◽  
Ehab N. M. Jirgis ◽  
Ngoc Anh Thu Ho ◽  
Tamsyn Stanborough ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Active Site ◽  
Feedback Inhibition ◽  
Feedback Regulation ◽  
Signal Propagation ◽  
Biosynthesis Pathway ◽  
Human Pathogen ◽  
Energy Status ◽  
Allosteric Site ◽  
Arginine Residues

Menaquinone (vitamin K2) plays a vital role in energy generation and environmental adaptation in many bacteria, including the human pathogen Mycobacterium tuberculosis (Mtb). Although menaquinone levels are known to be tightly linked to the cellular redox/energy status of the cell, the regulatory mechanisms underpinning this phenomenon are unclear. The first committed step in menaquinone biosynthesis is catalyzed by MenD, a thiamine diphosphate–dependent enzyme comprising three domains. Domains I and III form the MenD active site, but no function has yet been ascribed to domain II. Here, we show that the last cytosolic metabolite in the menaquinone biosynthesis pathway, 1,4-dihydroxy-2-naphthoic acid (DHNA), binds to domain II of Mtb-MenD and inhibits its activity. Using X-ray crystallography of four apo- and cofactor-bound Mtb-MenD structures, along with several spectroscopy assays, we identified three arginine residues (Arg-97, Arg-277, and Arg-303) that are important for both enzyme activity and the feedback inhibition by DHNA. Among these residues, Arg-277 appeared to be particularly important for signal propagation from the allosteric site to the active site. This is the first evidence of feedback regulation of the menaquinone biosynthesis pathway in bacteria, identifying a protein-level regulatory mechanism that controls menaquinone levels within the cell and may therefore represent a good target for disrupting menaquinone biosynthesis in M. tuberculosis.

New 9-methyl-8-(4-hydroxyphenyl)adenine derivatives as A1 adenosine receptor antagonists

2011 ◽  
Vol 76 (11) ◽  
pp. 1379-1393 ◽  
Author(s):  
Catia Lambertucci ◽  
Michela Buccioni ◽  
Barbara Cacciari ◽  
Diego Dal Ben ◽  
Stephanie Federico ◽  
...  
Keyword(s):  
Adenosine Receptor ◽  
Binding Assay ◽  
Radioligand Binding ◽  
Functional Assay ◽  
Receptor Subtypes ◽  
A1 Adenosine Receptor ◽  
Nanomolar Range ◽  
Mouse Ileum ◽  
Good Agreement ◽  
Adenine Derivatives

A new series of 9-methyladenines, bearing different bulky groups at the 8-position, were prepared and their affinity for the four human adenosine receptor subtypes were evaluated. All the synthesized compounds showed affinities at the A1, A2A, and A3AR subtypes ranging from nanomolar to micromolar levels with different degrees of A1selectivity, while they resulted nearly inactive at A2BAR. In particular, 9-methyl-8-[4-(4-methylbenzyloxy)phenyl]- adenine showed A1AR affinity in the nanomolar range and good levels of selectivity versus the other receptor subtypes. Furthermore, a functional assay at mouse ileum allowed to assess the potency of selected compounds at A1AR subtype. Results showed that all the tested derivatives are neutral antagonists and theirKbvalues are in good agreement with theKivalues from radioligand binding assay at human A1AR, confirming that the effect is due to inhibition of this subtype.

scholarly journals Comparison of a New Multiplex Binding Assay versus the Enzyme-Linked Immunosorbent Assay for Measurement of Serotype-Specific Pneumococcal Capsular Polysaccharide IgG

2011 ◽  
Vol 18 (10) ◽  
pp. 1744-1751 ◽  
Author(s):  
David Goldblatt ◽  
Lindsey Ashton ◽  
Yuhua Zhang ◽  
Joseph Antonello ◽  
Rocio D. Marchese
Keyword(s):  
Immunosorbent Assay ◽  
Simultaneous Measurement ◽  
Capsular Polysaccharide ◽  
Binding Assay ◽  
Enzyme Linked Immunosorbent Assay ◽  
Meso Scale Discovery ◽  
New Methods ◽  
Detection Assay ◽  
Heptavalent Pneumococcal Conjugate Vaccine ◽  
Good Agreement

ABSTRACTThe measurement of serotype-specific anti-capsular polysaccharide antibodies remains the mainstay of pneumococcal (Pn) vaccine evaluation. New methods that allow the simultaneous measurement of antibodies to several antigens in small volumes of serum, and that agree well with existing techniques, are urgently required to support the increasing number of concomitant vaccines delivered in the infant immunization schedules and the use of extended-valency Pn vaccines. We therefore compared a relatively new multiplexed platform for measuring anti-Pn antibodies with the existing WHO consensus enzyme-linked immunosorbent assay (ELISA). A panel of 50 pediatric samples (34 collected after receipt of a heptavalent pneumococcal conjugate vaccine [PCV7] and 16 without PCV7) was analyzed across two different laboratories using a new multiplex electrochemiluminescence (ECL)-based detection assay developed for the quantitation of IgG serotype-specific antipneumococcal antibodies, and the results were compared to those obtained using the WHO consensus ELISA. For the seven serotypes measured, there was good agreement between the techniques and laboratories. The most notable difference was found between the ECL assay and the ELISA: concentrations tended to be higher in the ECL assay. For serotypes 6B, 9V, 18C, and 23F, the average increases in concentration ranged from 48 to 102%. However, the agreement rates on the proportions of samples with concentrations surrounding 0.35 μg/ml were >82% for all serotypes tested. Agreement between the two laboratories running the ECL assay was generally good: agreement on proportions of samples with concentrations surrounding 0.35 μg/ml was in excess of 92%, and agreement on average antibody concentrations was within 31%. We conclude that the Meso Scale Discovery (MSD) platform provides a promising new technique for the simultaneous measurement of antipneumococcal antibodies.

Regulation of the early GA biosynthesis pathway in pea

Planta ◽  
2005 ◽  
Vol 222 (6) ◽  
pp. 1010-1019 ◽  
Author(s):  
Sandra E. Davidson ◽  
Stephen M. Swain ◽  
James B. Reid
Keyword(s):  
Biosynthesis Pathway ◽  
Ga Biosynthesis
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