#METABOLOMICS WORKBENCH nchorna_20201127_034822 DATATRACK_ID:2331 STUDY_ID:ST001609 ANALYSIS_ID:AN002643 PROJECT_ID:PR001034 VERSION 1 CREATED_ON November 30, 2020, 11:40 am #PROJECT PR:PROJECT_TITLE Comparative metabolomics analysis of two Saccharomyces cerevisiae strains: the PR:PROJECT_TITLE wild type and mtl1Δ, which carries a deletion of the mechanosensor Mtl1p PR:PROJECT_SUMMARY To gather more in-depth knowledge of the Mtl1p mechanosensor's role in PR:PROJECT_SUMMARY Saccharomyces cerevisiae metabolism, we conducted a comparative metabolomic PR:PROJECT_SUMMARY analysis of two Saccharomyces cerevisiae strains: the wild type and mtl1Δ, PR:PROJECT_SUMMARY which carries a deletion of the mechanosensor Mtl1p. Both strains were grown PR:PROJECT_SUMMARY under normal conditions at 27°C. The most significant metabolic changes between PR:PROJECT_SUMMARY these strains were related to amino acid metabolism, purine metabolism, and PR:PROJECT_SUMMARY carboxylic acid metabolism. PR:INSTITUTE University of Puerto Rico, Medical Sciences Campus PR:DEPARTMENT Biochemistry PR:LAST_NAME Chorna PR:FIRST_NAME Nataliya PR:ADDRESS University of Puerto Rico, Medical Sciences Campus, Department of Biochemistry, PR:ADDRESS Main Building, 6th Floor, Room A-632, San Juan, PR 00935 PR:EMAIL nataliya.chorna@upr.edu PR:PHONE 787-758-2525 x 1640 PR:FUNDING_SOURCE NIGMS-NIH-PRINBRE-P20GM103475 PR:CONTRIBUTORS Nelson Martínez–Matías, Sahily González–Crespo #STUDY ST:STUDY_TITLE Comparative metabolomics analysis of two Saccharomyces cerevisiae strains: the ST:STUDY_TITLE wild type and mtl1Δ, which carries a deletion of the mechanosensor Mtl1p. ST:STUDY_SUMMARY To gather more in-depth knowledge of the Mtl1p mechanosensor's role in ST:STUDY_SUMMARY Saccharomyces cerevisiae metabolism, we conducted a comparative metabolomic ST:STUDY_SUMMARY analysis of two Saccharomyces cerevisiae strains: the wild type and mtl1Δ, ST:STUDY_SUMMARY which carries a deletion of the mechanosensor Mtl1p. Both strains were grown ST:STUDY_SUMMARY under normal conditions at 27°C. The most significant metabolic changes between ST:STUDY_SUMMARY these strains were related to amino acid metabolism, purine metabolism, and ST:STUDY_SUMMARY carboxylic acid metabolism. ST:INSTITUTE University of Puerto Rico, Medical Sciences Campus ST:LAST_NAME Chorna ST:FIRST_NAME Nataliya ST:ADDRESS University of Puerto Rico, Medical Sciences Campus, Department of Biochemistry, ST:ADDRESS Main Building, 6th Floor, Room A-632, San Juan, PR 00935 ST:EMAIL nataliya.chorna@upr.edu ST:PHONE 787-758-2525 ext 1640 ST:NUM_GROUPS 2 ST:TOTAL_SUBJECTS 26 #SUBJECT SU:SUBJECT_TYPE Yeast SU:SUBJECT_SPECIES Saccharomyces cerevisiae SU:TAXONOMY_ID 4932 SU:GENOTYPE_STRAIN BY4742, YGR023w SU:CELL_BIOSOURCE_OR_SUPPLIER Open Biosystems #SUBJECT_SAMPLE_FACTORS: SUBJECT(optional)[tab]SAMPLE[tab]FACTORS(NAME:VALUE pairs separated by |)[tab]Raw file names and additional sample data SUBJECT_SAMPLE_FACTORS - BY4742-1 Strain:Wild-type RAW_FILE_NAME=BY4742-1.qgd; OD600=0.625 SUBJECT_SAMPLE_FACTORS - BY4742-2 Strain:Wild-type RAW_FILE_NAME=BY4742-2.qgd; OD600=0.627 SUBJECT_SAMPLE_FACTORS - BY4742-3 Strain:Wild-type RAW_FILE_NAME=BY4742-3.qgd; OD600=0.594 SUBJECT_SAMPLE_FACTORS - BY4742-4 Strain:Wild-type RAW_FILE_NAME=BY4742-4.qgd; OD600=0.661 SUBJECT_SAMPLE_FACTORS - BY4742-5 Strain:Wild-type RAW_FILE_NAME=BY4742-5.qgd; OD600=0.689 SUBJECT_SAMPLE_FACTORS - BY4742-6 Strain:Wild-type RAW_FILE_NAME=BY4742-6.qgd; OD600=0.644 SUBJECT_SAMPLE_FACTORS - BY4742-7 Strain:Wild-type RAW_FILE_NAME=BY4742-7.qgd; OD600=0.667 SUBJECT_SAMPLE_FACTORS - BY4742-8 Strain:Wild-type RAW_FILE_NAME=BY4742-8.qgd; OD600=0.678 SUBJECT_SAMPLE_FACTORS - BY4742-9 Strain:Wild-type RAW_FILE_NAME=BY4742-9.qgd; OD600=0.660 SUBJECT_SAMPLE_FACTORS - BY4742-10 Strain:Wild-type RAW_FILE_NAME=BY4742-10.qgd; OD600=0.663 SUBJECT_SAMPLE_FACTORS - BY4742-11 Strain:Wild-type RAW_FILE_NAME=BY4742-11.qgd; OD600=0.682 SUBJECT_SAMPLE_FACTORS - BY4742-12 Strain:Wild-type RAW_FILE_NAME=BY4742-12.qgd; OD600=0.613 SUBJECT_SAMPLE_FACTORS - BY4742-13 Strain:Wild-type RAW_FILE_NAME=BY4742-13.qgd; OD600=0.683 SUBJECT_SAMPLE_FACTORS - YGR023w-1 Strain:mtl1Δ RAW_FILE_NAME=YGR023w-1.qgd; OD600=0.652 SUBJECT_SAMPLE_FACTORS - YGR023w-2 Strain:mtl1Δ RAW_FILE_NAME=YGR023w-2.qgd; OD600=0.627 SUBJECT_SAMPLE_FACTORS - YGR023w-3 Strain:mtl1Δ RAW_FILE_NAME=YGR023w-3.qgd; OD600=0.602 SUBJECT_SAMPLE_FACTORS - YGR023w-4 Strain:mtl1Δ RAW_FILE_NAME=YGR023w-4.qgd; OD600=0.661 SUBJECT_SAMPLE_FACTORS - YGR023w-5 Strain:mtl1Δ RAW_FILE_NAME=YGR023w-5.qgd; OD600=0.639 SUBJECT_SAMPLE_FACTORS - YGR023w-6 Strain:mtl1Δ RAW_FILE_NAME=YGR023w-6.qgd; OD600=0.713 SUBJECT_SAMPLE_FACTORS - YGR023w-7 Strain:mtl1Δ RAW_FILE_NAME=YGR023w-7.qgd; OD600=0.635 SUBJECT_SAMPLE_FACTORS - YGR023w-8 Strain:mtl1Δ RAW_FILE_NAME=YGR023w-8.qgd; OD600=0.694 SUBJECT_SAMPLE_FACTORS - YGR023w-9 Strain:mtl1Δ RAW_FILE_NAME=YGR023w-9.qgd; OD600=0.685 SUBJECT_SAMPLE_FACTORS - YGR023w-10 Strain:mtl1Δ RAW_FILE_NAME=YGR023w-10.qgd; OD600=0.628 SUBJECT_SAMPLE_FACTORS - YGR023w-11 Strain:mtl1Δ RAW_FILE_NAME=YGR023w-11.qgd; OD600=0.681 SUBJECT_SAMPLE_FACTORS - YGR023w-12 Strain:mtl1Δ RAW_FILE_NAME=YGR023w-12.qgd; OD600=0.684 SUBJECT_SAMPLE_FACTORS - YGR023w-13 Strain:mtl1Δ RAW_FILE_NAME=YGR023w-13.qgd; OD600=0.683 #COLLECTION CO:COLLECTION_SUMMARY Cells were grown at 27°C, 210 rpm in 5 mL of complete synthetic medium (CSM, CO:COLLECTION_SUMMARY 0.67% Nitrogen base without amino acids and ammonium sulfate, 2% glucose). The CO:COLLECTION_SUMMARY next day, cells were replaced with fresh CSM and grow at 27°C to reach an OD of CO:COLLECTION_SUMMARY 0.6 - 0.7. 25 ml of each sample were harvested by centrifugation at 3,838 x g CO:COLLECTION_SUMMARY for 5 min at 4°C, washed with 1 ml ice cold sterile water, quenched using ice CO:COLLECTION_SUMMARY cold methanol, and stored at -80°C. CO:SAMPLE_TYPE Yeast cells #TREATMENT TR:TREATMENT_SUMMARY No treatment #SAMPLEPREP SP:SAMPLEPREP_SUMMARY Extraction of metabolites was performed by homogenization in 1 mL of cold SP:SAMPLEPREP_SUMMARY methanol/H2O (1:1) extraction solution and centrifugated at 167 x g at 4°C for SP:SAMPLEPREP_SUMMARY 5 min. Supernatants were collected and evaporated to dryness in a nitrogen SP:SAMPLEPREP_SUMMARY stream stream at 50 ºC (RapidVap, Labconco). Nitrilation was performed by SP:SAMPLEPREP_SUMMARY adding 150 μL of 0.2 mM hydroxylammonium chloride in pyridine to the dried SP:SAMPLEPREP_SUMMARY sample, and then heating at 90ºC for 40 minutes. After that, acetylation was SP:SAMPLEPREP_SUMMARY performed by adding 250 μL of acetic anhydride, and then heating at 90 ºC for SP:SAMPLEPREP_SUMMARY 60 minutes. After that, the sample was dried again in a nitrogen gas stream, and SP:SAMPLEPREP_SUMMARY then redissolved in 400 μL of ethyl acetate. Derivatized samples were collected SP:SAMPLEPREP_SUMMARY and stored at -20ºC. #CHROMATOGRAPHY CH:CHROMATOGRAPHY_TYPE GC CH:INSTRUMENT_NAME Shimadzu GCMS-QP2010 ultra CH:COLUMN_NAME Shimadzu SH-RXI (30M x 0.25mm x 0.25um) #ANALYSIS AN:ANALYSIS_TYPE MS #MS MS:INSTRUMENT_NAME Shimadzu QP2010 Ultra MS:INSTRUMENT_TYPE Single quadrupole MS:MS_TYPE EI MS:ION_MODE POSITIVE MS:MS_COMMENTS Inlet temperature was 280°C; ion source temperature was 200 °C. MS MS:MS_COMMENTS conditions were set as follows: SIM acquisition mode, electron energy of MS:MS_COMMENTS 70 eV, quadrupole scan range of m/z 115; 187; 212;314. Data were processed MS:MS_COMMENTS using the GCMS Solution Postrun Analysis software (Shimadzu Inc) for metabolites MS:MS_COMMENTS identification from their electron impact mass spectra by comparison to the NIST MS:MS_COMMENTS 2014 spectral mass library. #MS_METABOLITE_DATA MS_METABOLITE_DATA:UNITS mM MS_METABOLITE_DATA_START Samples BY4742-1 BY4742-2 BY4742-3 BY4742-4 BY4742-5 BY4742-6 BY4742-7 BY4742-8 BY4742-9 BY4742-10 BY4742-11 BY4742-12 BY4742-13 YGR023w-1 YGR023w-2 YGR023w-3 YGR023w-4 YGR023w-5 YGR023w-6 YGR023w-7 YGR023w-8 YGR023w-9 YGR023w-10 YGR023w-11 YGR023w-12 YGR023w-13 Factors Strain:Wild-type Strain:Wild-type Strain:Wild-type Strain:Wild-type Strain:Wild-type Strain:Wild-type Strain:Wild-type Strain:Wild-type Strain:Wild-type Strain:Wild-type Strain:Wild-type Strain:Wild-type Strain:Wild-type Strain:mtl1Δ Strain:mtl1Δ Strain:mtl1Δ Strain:mtl1Δ Strain:mtl1Δ Strain:mtl1Δ Strain:mtl1Δ Strain:mtl1Δ Strain:mtl1Δ Strain:mtl1Δ Strain:mtl1Δ Strain:mtl1Δ Strain:mtl1Δ Glucose 0.038140 0.039380 0.060200 0.050120 0.069050 0.041340 0.042350 0.037060 0.085690 0.038010 0.051820 0.050020 0.044300 0.12715 0.06061 0.04862 0.06086 0.03731 0.03711 0.03789 0.03592 0.04124 0.0383 0.0839 0.04703 0.05165 MS_METABOLITE_DATA_END #METABOLITES METABOLITES_START metabolite_name quantified m/z KEGG Glucose 180 C00031 METABOLITES_END #END