Summary of Study ST000817
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 PR000582. The data can be accessed directly via it's Project DOI: 10.21228/M8FD4K This work is supported by NIH grant, U2C- DK119886.
See: https://www.metabolomicsworkbench.org/about/howtocite.php
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.
| Study ID | ST000817 |
| Study Title | Dynamics of metabolism, proliferation and differentiation in Toxoplasma gondii mutants deficient in glycolysis and gluconeogenesis |
| Study Type | Time course 13C labeling of Toxoplasma gondii using glucose and glutamine |
| Study Summary | The focus of this study was to profile the metabolic fates of glucose and glutamine in wild type, glycolytic mutant (hexokinase KO) and gluconeogenesis mutant (PEPCK1 KO) tachyzoite stage T. gondii parasites. 13C6-glucose and 13C5-15N1-glutamine were used as metabolic tracers. Metabolites from glycolysis, pentosephosphate pathway and Kerbs cycle were identified and their isotopomer abundance was quantified in order to visualize metabolic flux across the indicated pathways. A time course (from 0 minutes to 120 minutes) analysis was carried out in the labeling experiments. Here, host cell free extracellular tachyzoite stage T. gondii parasites were used. |
| Institute | CSIR-National Chemical Laboratory |
| Department | Biochemical Sciences Division |
| Laboratory | Molecular Parasitology Laboratory |
| Last Name | Shanmugam |
| First Name | Dhanasekaran |
| Address | Dr. Homi Bhabha Road, Pune, 411008, Maharashtra, India |
| d.shanmugam@ncl.res.in | |
| Phone | +91-20-25902719 |
| Submit Date | 2017-07-22 |
| Raw Data Available | Yes |
| Raw Data File Type(s) | mzXML |
| Analysis Type Detail | LC-MS |
| Release Date | 2017-10-11 |
| Release Version | 1 |
Select appropriate tab below to view additional metadata details:
Sample Preparation:
| Sampleprep ID: | SP000850 |
| Sampleprep Summary: | Metabolic labeling, metabolite extraction, and LC-MS profiling – DMEM minus glucose media supplemented with +613C glucose (5.5 mM) and MEM minus glutamine media supplemented with +513C-+213N glutamine (4 mM) were used as culture mediums for metabolic labeling studies using RH wt, RH ∆ku80, RH ∆hk, and RH ∆pepck1 parasites. Freshly isolated extracellular (host cell free) tachyzoites stage parasites were washed once in complete medium and resuspended in either 13C labeled glucose or glutamine (Cambridge Isotope Laboratories, Inc.) containing medium at a density of 108 parasites per milliliter. Labeling was allowed to proceed over a time period of 5, 10, 15, 30, 60 and 120 minutes in 1 ml culture suspension. At each time point, metabolites were extracted from replicate parasite samples using a modified version of a previously reported protocol. Briefly, host cell free parasites were collected by centrifugation at 3000 rpm for 5 minutes in cold and then immediately resuspended in 200 µls of ice cold 80% acetonitrile (Chem-Impex; JT Bakers) in water and incubated in ice for 15 minutes with intermittent vortexing. The supernatant was then collected after centrifuging at 13,000 rpm for 5 minutes and the pellet was further extracted twice with 100 µls of the same solvent using ultrasound in a sonicating iced water bath for 15 minutes. All the extracts were pooled (total 400 µls) and stored in -80°C until further processing for liquid chromatography coupled mass spectrometry (LC-MS) analysis. |
| Extraction Method: | Hydrophilic organic extract |
| Extract Storage: | -80 deg. Cel. until LC-MS analysis is done |
| Sample Resuspension: | The acetonitrile:water extracts from parasites were dried under nitrogen flow and resuspended in 200 µls of water:methanol (97:3) containing 10 mM tributylamine and 15 mM acetic acid. |
| Sample Derivatization: | n/a |
| Sample Spiking: | PIPES @ 100ng/ml final concentration |
