Summary of Study ST002232

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 PR001422. The data can be accessed directly via it's Project DOI: 10.21228/M8X411 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 IDST002232
Study TitleSteady-state metabolomics Saccharomyces cerevisiae mitochondrial fatty acid synthesis (mtFAS) mutants and CTP1 overexpression
Study TypeSteady-state targeted and untargeted metabolomics
Study SummaryThe goal of this work was to analyze metabolic changes in yeast with the mct1 gene knock-out or CTP1 overexpression conditions using liquid chromatography-mass spectrometry (LC-MS).
Institute
University of Utah
DepartmentBiochemistry
LaboratoryRutter
Last NameBerg
First NameJordan
Address15 N Medical Drive East RM 5520, Salt Lake City, UT 84112-5650 USA
Emailjordan.berg@biochem.utah.edu
Phone+1 (801) 581 3340
Submit Date2022-07-10
Raw Data AvailableYes
Raw Data File Type(s)mzXML
Analysis Type DetailOther
Release Date2022-08-08
Release Version1
Jordan Berg Jordan Berg
https://dx.doi.org/10.21228/M8X411
ftp://www.metabolomicsworkbench.org/Studies/ application/zip

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Project:

Project ID:PR001422
Project DOI:doi: 10.21228/M8X411
Project Title:Steady-state metabolomics Saccharomyces cerevisiae mitochondrial fatty acid synthesis (mtFAS) mutants and CTP1 overexpression
Project Type:Steady-state targeted and untargeted metabolomics
Project Summary:The goal of this work was to analyze metabolic changes in yeast with the mct1 gene knock-out or CTP1 overexpression conditions using liquid chromatography-mass spectrometry (LC-MS).
Institute:University of Utah
Department:Biochemistry
Laboratory:Rutter
Last Name:Berg
First Name:Jordan
Address:15 N Medical Drive East RM 5520, Salt Lake City, UT 84112-5650 USA
Email:jordan.berg@biochem.utah.edu
Phone:+1 (801) 581 3340
Funding Source:R35GM131854
Publications:https://doi.org/10.1101/2020.06.25.171850
Contributors:Yeyun Ouyang, Ahmad Cluntun, Jordan Berg

Subject:

Subject ID:SU002318
Subject Type:Yeast
Subject Species:Saccharomyces cerevisiae
Taxonomy ID:4932
Genotype Strain:BY4743 (MATa/\textalpha, his3/his3, leu2/leu2, ura3/ura3, met15/MET15, lys2/LYS2)

Factors:

Subject type: Yeast; Subject species: Saccharomyces cerevisiae (Factor headings shown in green)

mb_sample_id local_sample_id Strain Genotype Experimental Factor 1 - Vector Experimental Factor 2 - Media Experimental Factor 2 - Replicate
SA212639MCT_A1_SDBY4743 mct1 deletion CTP1-overexpression vector S-complete + 2% glucose replicate 1
SA212640MCT_A2_SDBY4743 mct1 deletion CTP1-overexpression vector S-complete + 2% glucose replicate 2
SA212641MCT_A3_SDBY4743 mct1 deletion CTP1-overexpression vector S-complete + 2% glucose replicate 3
SA212642MCT_A1_SRBY4743 mct1 deletion CTP1-overexpression vector S-complete + 2% raffinose replicate 1
SA212643MCT_A2_SRBY4743 mct1 deletion CTP1-overexpression vector S-complete + 2% raffinose replicate 2
SA212644MCT_A3_SRBY4743 mct1 deletion CTP1-overexpression vector S-complete + 2% raffinose replicate 3
SA212645MCT_EV1_SDBY4743 mct1 deletion empty vector S-complete + 2% glucose replicate 1
SA212646MCT_EV2_SDBY4743 mct1 deletion empty vector S-complete + 2% glucose replicate 2
SA212647MCT_EV3_SDBY4743 mct1 deletion empty vector S-complete + 2% glucose replicate 3
SA212648MCT_EV1_SRBY4743 mct1 deletion empty vector S-complete + 2% raffinose replicate 1
SA212649MCT_EV2_SRBY4743 mct1 deletion empty vector S-complete + 2% raffinose replicate 2
SA212650MCT_EV3_SRBY4743 mct1 deletion empty vector S-complete + 2% raffinose replicate 3
SA212651WT_A1_SDBY4743 wild type CTP1-overexpression vector S-complete + 2% glucose replicate 1
SA212652WT_A2_SDBY4743 wild type CTP1-overexpression vector S-complete + 2% glucose replicate 2
SA212653WT_A3_SDBY4743 wild type CTP1-overexpression vector S-complete + 2% glucose replicate 3
SA212654WT_A1_SRBY4743 wild type CTP1-overexpression vector S-complete + 2% raffinose replicate 1
SA212655WT_A2_SRBY4743 wild type CTP1-overexpression vector S-complete + 2% raffinose replicate 2
SA212656WT_A3_SRBY4743 wild type CTP1-overexpression vector S-complete + 2% raffinose replicate 3
SA212657WT_EV1_SDBY4743 wild type empty vector S-complete + 2% glucose replicate 1
SA212658WT_EV2_SDBY4743 wild type empty vector S-complete + 2% glucose replicate 2
SA212659WT_EV3_SDBY4743 wild type empty vector S-complete + 2% glucose replicate 3
SA212660WT_EV1_SRBY4743 wild type empty vector S-complete + 2% raffinose replicate 1
SA212661WT_EV2_SRBY4743 wild type empty vector S-complete + 2% raffinose replicate 2
SA212662WT_EV3_SRBY4743 wild type empty vector S-complete + 2% raffinose replicate 3
Showing results 1 to 24 of 24

Collection:

Collection ID:CO002311
Collection Summary:Metabolomics data were generated by growing the appropriate yeast strains in synthetic complete media supplemented with 2% glucose until they reached saturation (n=3). Cells were then transferred to S-minimal media containing 2% raffinose and leucine and harvested after approximately 8 hours (n=3) at OD600=0.6-0.8. The procedures for metabolite extraction were performed as previously described in [Bricker et al., Science, 2012]. Yeast cultures were pelleted, snap-frozen and kept at −80°C. 5ml of 75% boiled ethanol was added to every frozen pellet. Pellets were vortexed and incubated at 90°C for 5 minutes. All samples were then centrifuged at 5,000 Relative Centrifugal Force (RCF) for 10 minutes. Supernatants were transferred to fresh tubes, evaporated overnight in a Speed Vacuum, and then stored at −80°C until they were run on the mass spectrometer.
Sample Type:Yeast cells

Treatment:

Treatment ID:TR002330
Treatment Summary:N/A

Sample Preparation:

Sampleprep ID:SP002324
Sampleprep Summary:The conditions for liquid chromatography are described in previous studies [Cluntun et al., Cancer Metab., 2015; Lukey et al., Cell Rep., 2019]. Briefly, a hydrophilic interaction liquid chromatography method (HILIC) with an Xbridge amide column (100 × 2.1 mm, 3.5 μm) (Waters) was employed on a Dionex (Ultimate 3000 UHPLC) for compound separation and detection at room temperature. The mobile phase A was 20 mM ammonium acetate and 15 mM ammonium hydroxide in water with 3% acetonitrile, pH 9.0, and the mobile phase B was acetonitrile. The linear gradient was as follows: 0 min, 85% B; 1.5 min, 85% B, 5.5 min, 35% B; 10 min, 35% B, 10.5 min, 35% B, 14.5 min, 35% B, 15 min, 85% B, and 20 min, 85% B. The flow rate was 0.15 ml/min from 0 to 10 min and 15 to 20 min, and 0.3 ml/min from 10.5 to 14.5 min. All solvents were LC-MS grade and purchased from Thermo Fisher Scientific. Mass spectrometry was performed as described in previous studies [Cluntun et al., Cancer Metab., 2015; Lukey et al., Cell Rep., 2019]. Briefly, the Q Exactive MS (Thermo Scientific) is equipped with a heated electrospray ionization probe (HESI), and the relevant parameters are as listed: evaporation temperature, 120°C; sheath gas, 30; auxiliary gas, 10; sweep gas, 3; spray voltage, 3.6 kV for positive mode and 2.5 kV for negative mode. Capillary temperature was set at 320°C, and S-lens was 55. A full scan range from 60 to 900 (m/z) was used. The resolution was set at 70,000. The maximum injection time was 200 ms. Automated gain control (AGC) was targeted at 3,000,000 ions. Data were collected, metabolites were identified, and their peak area was recorded using El-MAVEN software [Agrawal et al., Methods Mol. Biol., 2019; Clasquin et al., Curr. Protoc. Bioinformatics, 2012; Melamud et al., Anal. Chem., 2010]. These data were transferred to an Excel spreadsheet (Microsoft, Redmond WA). Metabolite identity was established using a combination of an in-house metabolite library developed using pure purchased standards, the NIST (https://www.nist.gov) and Fiehn libraries [Kind et al, Anal. Chem., 2009]. P-values were derived using a homoscedastic, two-tailed Student's T-test and adjusted using the Benjamini-Hochberg correction procedure.

Combined analysis:

Analysis ID AN003642
Analysis type MS
Chromatography type HILIC
Chromatography system Thermo Dionex Ultimate 3000
Column Waters XBridge Amide (100 x 4.6mm, 3.5um)
MS Type Other
MS instrument type LC-Q Exactive
MS instrument name Q Exactive
Ion Mode UNSPECIFIED
Units peak height

Chromatography:

Chromatography ID:CH002696
Chromatography Summary:The conditions for liquid chromatography are described in previous studies [Cluntun et al., Cancer Metab., 2015; Lukey et al., Cell Rep., 2019]. Briefly, a hydrophilic interaction liquid chromatography method (HILIC) with an Xbridge amide column (100 × 2.1 mm, 3.5 μm) (Waters) was employed on a Dionex (Ultimate 3000 UHPLC) for compound separation and detection at room temperature. The mobile phase A was 20 mM ammonium acetate and 15 mM ammonium hydroxide in water with 3% acetonitrile, pH 9.0, and the mobile phase B was acetonitrile. The linear gradient was as follows: 0 min, 85% B; 1.5 min, 85% B, 5.5 min, 35% B; 10 min, 35% B, 10.5 min, 35% B, 14.5 min, 35% B, 15 min, 85% B, and 20 min, 85% B. The flow rate was 0.15 ml/min from 0 to 10 min and 15 to 20 min, and 0.3 ml/min from 10.5 to 14.5 min. All solvents were LC-MS grade and purchased from Thermo Fisher Scientific.
Instrument Name:Thermo Dionex Ultimate 3000
Column Name:Waters XBridge Amide (100 x 4.6mm, 3.5um)
Flow Gradient:0 min, 85% B; 1.5 min, 85% B, 5.5 min, 35% B; 10 min, 35% B, 10.5 min, 35% B, 14.5 min, 35% B, 15 min, 85% B, and 20 min, 85% B
Flow Rate:0.15 ml/min from 0 to 10 min and 15 to 20 min, and 0.3 ml/min from 10.5 to 14.5 min
Solvent A:20 mM ammonium acetate and 15 mM ammonium hydroxide in water with 3% acetonitrile, pH 9.0
Solvent B:acetonitrile
Chromatography Type:HILIC

MS:

MS ID:MS003393
Analysis ID:AN003642
Instrument Name:Q Exactive
Instrument Type:LC-Q Exactive
MS Type:Other
MS Comments:Data were collected, metabolites were identified, and their peak area was recorded using El-MAVEN software [Agrawal et al., Methods Mol. Biol., 2019; Clasquin et al., Curr. Protoc. Bioinformatics, 2012; Melamud et al., Anal. Chem., 2010]. These data were transferred to an Excel spreadsheet (Microsoft, Redmond WA). Metabolite identity was established using a combination of an in-house metabolite library developed using pure purchased standards, the NIST (https://www.nist.gov) and Fiehn libraries [Kind et al, Anal. Chem., 2009]. P-values were derived using a homoscedastic, two-tailed Student's T-test and adjusted using the Benjamini-Hochberg correction procedure.
Ion Mode:UNSPECIFIED
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