Summary of Study ST002810

This data is available at the NIH Common Fund's National Metabolomics Data Repository (NMDR) website, the Metabolomics Workbench, https://www.metabolomicsworkbench.org, where it has been assigned Project ID PR001757. The data can be accessed directly via it's Project DOI: 10.21228/M8M13Z This work is supported by NIH grant, U2C- DK119886.

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This study contains a large results data set and is not available in the mwTab file. It is only available for download via FTP as data file(s) here.

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Study IDST002810
Study TitleGAS2 encodes a 2-oxoglutarate dependent dioxygenase involved in ABA catabolism
Study SummaryLiu et al. [1] recently reported the characterization of Arabidopsis thaliana GAS2 (Gain of Function in ABA-modulated Seed Germination 2), which was described as an enzyme that catalyzes the stereospecific hydration of GA12 to produce GA12 16, 17-dihydro-16α-ol (DHGA12). A second paper describes the conversion of GA12 to an unidentified product by GAS2 and also reports that this enzyme does not convert ABA [2]. However, as previously reported [3], we did not find any conversion of [17-14C]-labeled or [1-,7-,12-,18-14C4]-labeled GA12 by GAS2. Instead, we present here data showing that the recombinant GAS2 enzyme is capable of catabolising abscisic acid (ABA) to phaseic acid (PA) and further to a second product, putative 8’-carboxy-ABA (compound A; Fig. 1a) [4]. References: [1] Liu, H. et al. Biosynthesis of DHGA12 and its roles in Arabidopsis seedling establishment. Nat. Commun. 10, 1768 (2019). [2] Xiong, W. et al. The dioxygenase GIM2 functions in seed germination by altering gibberellin production in Arabidopsis. J. Integr. Plant Biol. 60, 276-291 (2018). [3] Lange, T. & Pimenta Lange, M. J. The Multifunctional Dioxygenases of Gibberellin Synthesis. Plant Cell Physiol. 61, 1869-1879 (2020). [4] Lange, T., Atiq, N., Pimenta Lange. GAS2 encodes a 2-oxoglutarate dependent dioxygenase involved in ABA catabolism. bioRxiv, doi: 10.1101/2022.11.16.516706 (2022).
Institute
Technische Universität Braunschweig
DepartmentBiochemie und Physiology der Pflanzen
LaboratoryAG Lange
Last NameLange
First NameTheo
AddressMendelssohnstr. 4
Emailtheo.lange@tu-bs.de
Phone00495313915880
Submit Date2023-08-01
Raw Data AvailableYes
Raw Data File Type(s)raw(Thermo)
Analysis Type DetailGC-MS
Release Date2023-10-11
Release Version1
Theo Lange Theo Lange
https://dx.doi.org/10.21228/M8M13Z
ftp://www.metabolomicsworkbench.org/Studies/ application/zip

Select appropriate tab below to view additional metadata details:

Project:

Project ID:PR001757
Project DOI:doi: 10.21228/M8M13Z
Project Title:Functional GAS2 studies
Project Type:GC-MS qualitative analysis
Project Summary:Liu et al. recently reported the characterization of Arabidopsis thaliana GAS2 (Gain of Function in ABA-modulated Seed Germination 2), which was described as an enzyme that catalyzes the stereospeci?c hydration of GA12 to produce GA12 16, 17-dihydro-16?-ol (DHGA12). A second paper describes the conversion of GA12 to an unidentified product by GAS2 and also reports that this enzyme does not convert ABA. However, as previously reported, we did not find any conversion of [17-14C]-labeled or [1-,7-,12-,18-14C4]-labeled GA12 by GAS2. Instead, we present here data showing that the recombinant GAS2 enzyme is capable of catabolising abscisic acid (ABA) to phaseic acid (PA) and further to a second product, putative 8’-carboxy-ABA (compound A; Fig. 1a).
Institute:Technische Universität Braunschweig
Department:Biochemie und Physiologie der Pflanzen
Laboratory:AG Lange
Last Name:Lange
First Name:Theo
Address:Mendelssohnstr. 4
Email:theo.lange@tu-bs.de
Phone:00495313915880
Contributors:Nadiem Atiq, Maria João Pimenta Lange

Subject:

Subject ID:SU002917
Subject Type:Plant
Subject Species:Arabidopsis thaliana

Factors:

Subject type: Plant; Subject species: Arabidopsis thaliana (Factor headings shown in green)

mb_sample_id local_sample_id Treatment
SA301609551_1020ox1
SA301610552_1120ox1
SA301611550_0920ox1
SA301612559_03ABA
SA301642539_06d3-PA
SA301643868_08d6-ABA
SA301613266_01DHGA12
SA301614920_10GAS2
SA301615967_01GAS2
SA301616907_09GAS2
SA301617969_02GAS2
SA301618748_05GAS2
SA301619747_04GAS2
SA301620970_03GAS2
SA301621990_02GAS2
SA301622403_02GAS2
SA301623976_07GAS2
SA301624975_06GAS2
SA301625972_04GAS2
SA301626974_05GAS2
SA301627746_03GAS2
SA301628691_02GAS2
SA301629543_08GAS2
SA301630481_03GAS2
SA301631542_07GAS2
SA301632486_06GAS2
SA301633485_05GAS2
SA301634482_04GAS2
SA301635480_02GAS2
SA301636583_04GAS2
SA301637585_05GAS2
SA301638479_01GAS2
SA301639556_02GAS2
SA301640558_07GAS2
SA301641538_01PA
Showing results 1 to 35 of 35

Collection:

Collection ID:CO002910
Collection Summary:Recombinant GAS2 and AtGA20ox1 were produced as previously described [5,6]. [5] Pimenta Lange, M. J. et al. Functional characterization of gibberellin oxidases from cucumber, Cucumis sativus L. Phytochemistry 90, 62-69 (2013). [6] Pimenta Lange, M. J. et al. Cucumber gibberellin 1-oxidase/desaturase initiates novel gibberellin catabolic pathways. J. Biol. Chem. 295, 8442-8448 (2020).
Sample Type:Bacterial cells

Treatment:

Treatment ID:TR002926
Treatment Summary:Recombinant GAS2 and AtGA20ox1 were produced as previously described [5,6]. [5] Pimenta Lange, M. J. et al. Functional characterization of gibberellin oxidases from cucumber, Cucumis sativus L. Phytochemistry 90, 62-69 (2013). [6] Pimenta Lange, M. J. et al. Cucumber gibberellin 1-oxidase/desaturase initiates novel gibberellin catabolic pathways. J. Biol. Chem. 295, 8442-8448 (2020).

Sample Preparation:

Sampleprep ID:SP002923
Sampleprep Summary:3’,5’,5’,7’,7’,7’-d6-labelled ABA and 17,17-d2-labelled GA12 were purchased from OlChemIm, Czech Republic. PA and 7’,7’,7’-d3-PA were gifts from Professor Eiji Nambara (University of Toronto, Canada). Preparations of E. coli cell lysates were incubated in a total volume of 100 µl containing 100 mM Tris-HCl, pH 7.0 at 30°C for 16 h with 2-oxoglutarate and ascorbate (100mM each, final concentrations), FeSO4 (0.5 mM), catalase (1mg/ml), and the substrates (5 µl in methanol for ABA, 3’,5’,5’,7’,7’,7’-d6-labeled ABA (500 ng), PA (500 ng), and 7’,7’,7’-d3-labeled PA (50 ng), and 2 µl in methanol per 5 ng 17,17-d2-labelled GA12. Variations to the standard incubation conditions are indicated in the individual experiments. Incubation products were extracted and analyzed by reverse-phase HPLC as described previously12 using gradients of increasing methanol in water, containing 1% acetic acid, at 1 ml.min-1 as follows: 50% methanol, followed by five 0.5-min steps to 57.4%, 60.6%, 61.8%, 62.3%, 62.5%, one 5-min step to 62.7%, one 1-min step to 63.2% and seven 2-min steps to 64.3%, 67.2%, 70.5%, 74.9%, 81%, 89% and 100% methanol. PA and compound A eluted between 3 and 6 min, and ABA between 6 and 9 min. The dried HPLC-fractions were redissolved in 100 µl methanol and methylated with 100 µl ethereal diazomethane.

Combined analysis:

Analysis ID AN004570
Analysis type MS
Chromatography type GC
Chromatography system Thermo Scientific Trace 1300
Column Agilent DB5-MS (30m x 0.25mm, 0.25um)
MS Type EI
MS instrument type Single quadrupole
MS instrument name Thermo Scientific ISQ 7000
Ion Mode POSITIVE
Units Peak area

Chromatography:

Chromatography ID:CH003435
Instrument Name:Thermo Scientific Trace 1300
Column Name:Agilent DB5-MS (30m x 0.25mm, 0.25um)
Column Temperature:60-280
Flow Gradient:60-280
Flow Rate:1.2 ml/min
Solvent A:Helium
Solvent B:Helium
Chromatography Type:GC

MS:

MS ID:MS004316
Analysis ID:AN004570
Instrument Name:Thermo Scientific ISQ 7000
Instrument Type:Single quadrupole
MS Type:EI
MS Comments:ThermoScientific ISQ7000 with Advanced Electron Ionization (AEI) source. Software used was Chromeleon 7.2.10
Ion Mode:POSITIVE
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