Summary of Study ST003933
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 PR002463. The data can be accessed directly via it's Project DOI: 10.21228/M88K07 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 | ST003933 |
| Study Title | Lipidomic analysis of genetically engineered PCC 7002 indicate lipid remodeling and differential fatty acid production at varied temperatures |
| Study Summary | An untargeted lipidomics MS workflow was used to investigate the alterations of lipid profiles for two strains of gentetically modified PCC 7002 at two different temperatures. The two strains evaluated were WT and an aas-knockout strain. The temperatures at which both strains were subjected were 30°C and 37°C. In the Δaas mutant, which lacks an acyl-activating synthetase (aas) and secretes FFAs, namely C16:1-ACP, C18:1-ACP, C18:2-ACP, and C18:3-ACP, with those FFAs being accumulated at both optimal and cold temperatures. This was unexpected, as cyanobacteria typically do not produce polyunsaturated acyl-ACPs, suggesting compensatory acyltransferase or other enzyme activities that were further explored with proteomic analysis. Several candidates were identified that could potentially reactivate FFAs and would limit secretion that is an important aspect of current biotechnological approaches. |
| Institute | Vanderbilt University |
| Last Name | Shepard |
| First Name | Hawkins |
| Address | 7330 Stevenson Center, Station B 351822, Nashville, TN 37235 |
| hawkins.s.shepard@vanderbilt.edu | |
| Phone | 6153434563 |
| Submit Date | 2025-05-22 |
| Raw Data Available | Yes |
| Raw Data File Type(s) | mzML, d |
| Analysis Type Detail | LC-MS |
| Release Date | 2025-11-10 |
| Release Version | 1 |
Select appropriate tab below to view additional metadata details:
Project:
| Project ID: | PR002463 |
| Project DOI: | doi: 10.21228/M88K07 |
| Project Title: | Lipidomic analysis of genetically engineered PCC 7002 indicate lipid remodeling and differential fatty acid production at varied temperatures |
| Project Summary: | E. coli and cycanobacteria, including Picosynechococcus sp. PCC 7002 (PCC 7002) are promising hosts for biomanufacturing due to their rapid growth and genetic manipulability. Microbial production of free fatty acids provides a potential renewable source of acyl chains, but genetic engineering for high production strains remains challenging. Using an untargeted lipidomics mass spectrometry workflow, we investigated two strains of PCC 7002 grown at two temperatures in order to probe the alterations in lipid profiles associated with the different genetic and environmental conditions. |
| Institute: | Vanderbilt University |
| Last Name: | Shepard |
| First Name: | Hawkins |
| Address: | 7330 Stevenson Center, Station B 351822, Nashville, TN 37235 |
| Email: | hawkins.s.shepard@vanderbilt.edu |
| Phone: | 6153434563 |
Subject:
| Subject ID: | SU004069 |
| Subject Type: | Bacteria |
| Subject Species: | Picosynechococcus sp. PCC 7002 |
| Taxonomy ID: | 32049 |
Factors:
Subject type: Bacteria; Subject species: Picosynechococcus sp. PCC 7002 (Factor headings shown in green)
| mb_sample_id | local_sample_id | Sample source | Genotype | Treatment |
|---|---|---|---|---|
| SA446393 | aas37b | Cyanobacteria | aas-knockout | control |
| SA446394 | aas37d | Cyanobacteria | aas-knockout | control |
| SA446395 | aas37c | Cyanobacteria | aas-knockout | control |
| SA446396 | aas37a | Cyanobacteria | aas-knockout | control |
| SA446389 | aas30d | Cyanobacteria | aas-knockout | Low-tempeature |
| SA446390 | aas30c | Cyanobacteria | aas-knockout | Low-tempeature |
| SA446391 | aas30b | Cyanobacteria | aas-knockout | Low-tempeature |
| SA446392 | aas30a | Cyanobacteria | aas-knockout | Low-tempeature |
| SA446385 | WT37b | Cyanobacteria | Wild-type | control |
| SA446386 | WT37d | Cyanobacteria | Wild-type | control |
| SA446387 | WT37c | Cyanobacteria | Wild-type | control |
| SA446388 | WT37a | Cyanobacteria | Wild-type | control |
| SA446381 | WT30d | Cyanobacteria | Wild-type | Low-tempeature |
| SA446382 | WT30c | Cyanobacteria | Wild-type | Low-tempeature |
| SA446383 | WT30b | Cyanobacteria | Wild-type | Low-tempeature |
| SA446384 | WT30a | Cyanobacteria | Wild-type | Low-tempeature |
| Showing results 1 to 16 of 16 |
Collection:
| Collection ID: | CO004062 |
| Collection Summary: | The wild-type and aas-knockout strains of Picosynechococcus sp. PCC 7002 were cultivated in A+ media at 30°C and 37°C with shaking at 130 rpm, 1% carbon dioxide, 300 micromolar/square meter/second until reaching mid-log phase (OD730 of 0.6-0.7). 60 mL cultures were quenched with 90 mL of cold 30% MeOH/water (-20°C). Then the cell pellets were collected by centrifugation at 4,000g for 10 min at -4°C, the supernatant was decanted, and the pellets were snap frozen with liquid Nitrogen, lyophilized, and transferred into 2mL tubes and stored at -80°C until undergoing sample preparation |
| Sample Type: | Bacterial cells |
Treatment:
| Treatment ID: | TR004078 |
| Treatment Summary: | The two treatments/factors involved with this study were genotype and growth temperature. The two strains investigated were WT PCC 7002 and aas-knockout PCC 7002. Each strain was cultured at 30°C and 37°C. |
Sample Preparation:
| Sampleprep ID: | SP004075 |
| Sampleprep Summary: | The lyophilized Synechococcus sp. PCC 7002 pellets were resuspended in 2 mL of -20°C methanol. Portions of each sample were normalized based on OD730 measurements to a volume of 100 µL before adding 800 µL of cold MeOH. The samples were then incubated at -80°C overnight to facilitate protein precipitation. These samples were centrifuged for 10 minutes at 1000 x g (4 ˚C) before transferring the supernatant and drying it in vacuo. Dried samples were resuspended in 100 µL water, vortexed thoroughly, followed by 100 µL MeOH, and incubated at room temperature for 10 minutes. An isotopically labeled lipid mixture (SPLASH LIPIDOMIX, Avanti) was added for QA/QC purposes. A liquid-liquid extraction was performed by adding 800 µL MTBE, vortexing, and subsequently incubating on ice for 10 mins. After incubation, samples were centrifuged for 10 minutes at 1000 x g and 4°C. The nonpolar, MTBE layer containing the lipophilic components was removed and dried under vacuum. These fractions were resuspended for HPLC-MS analysis in 100 µL 50:50 IPA:MeOH containing 40 µg/mL heptadecanoic acid and nonadecanoic acid, as well as 10 µg/mL glucosyl(β) sphingosine and N-heptadecanoyl-D-erythrosphingosine. |
Chromatography:
| Chromatography ID: | CH004904 |
| Instrument Name: | Agilent 1290 Infinity |
| Column Name: | Thermo Hypersil GOLD AQ (100 x 2.1 mm, 1.9 µm) |
| Column Temperature: | 40°C |
| Flow Gradient: | 70-30% B in 0.5 min, 30-70% B in 1.5 min, 70-100% B in 13 min, 100% B for 6 min, 100-10% B in 1 min, 10% B for 2 min, 10-30% B in 1 min, and 30-70% B in 5 mins, followed by a 4-minute post-injection period for column equilibration |
| Flow Rate: | 0.25 mL/min |
| Solvent A: | 90% Water/10% Methanol; 10 mM Ammonium acetate |
| Solvent B: | 50% Isopropyl alcohol/30% Methanol/20% Acetonitrile |
| Chromatography Type: | Reversed phase |
Analysis:
| Analysis ID: | AN006458 |
| Analysis Type: | MS |
| Chromatography ID: | CH004904 |
| Has Mz: | 1 |
| Has Rt: | 1 |
| Rt Units: | Minutes |
| Results File: | ST003933_AN006458_Results.txt |
| Units: | m/z |
