#METABOLOMICS WORKBENCH huntermoseley_20190215_183041 DATATRACK_ID:1625 STUDY_ID:ST001139 ANALYSIS_ID:AN001869 VERSION 1 CREATED_ON 02-08-2024 #PROJECT PR:PROJECT_TITLE P4HA1 knockdown in the breast cell line MDA231 PR:PROJECT_SUMMARY Determine the effect of knocking down P4HA1 in the MDA231 cell line (including PR:PROJECT_SUMMARY alterations to metabolic pathways). PR:INSTITUTE University of Kentucky PR:DEPARTMENT Markey Cancer Center PR:LAST_NAME Xu PR:FIRST_NAME Ren PR:ADDRESS BBSRC361, 741 South Limestone, Lexington, KY 40536, USA PR:EMAIL ren.xu2010@uky.edu PR:PHONE 000-000-0000 PR:DOI http://dx.doi.org/10.21228/M8T10P #STUDY ST:STUDY_TITLE P4HA1 knockdown in the breast cell line MDA231 Gln metabolism (part VI) ST:STUDY_TYPE isotope tracer ST:STUDY_SUMMARY Determine the effect on glutamine metabolism of knocking down P4HA1 in the ST:STUDY_SUMMARY MDA231 cell line. ST:INSTITUTE University of Kentucky ST:DEPARTMENT Markey Cancer Center ST:LAST_NAME Xiong ST:FIRST_NAME Gaofeng ST:ADDRESS BBSRC361, 741 South Limestone, Lexington, KY 40536, USA ST:EMAIL gaofeng.xiong@uky.edu ST:PHONE 000-000-0000 ST:SUBMIT_DATE 2019-02-15 #SUBJECT SU:SUBJECT_TYPE Animal SU:SUBJECT_SPECIES Homo sapiens SU:TAXONOMY_ID 9606 #SUBJECT_SAMPLE_FACTORS: SUBJECT(optional)[tab]SAMPLE[tab]FACTORS(NAME:VALUE pairs separated by |)[tab]Additional sample data SUBJECT_SAMPLE_FACTORS - 2_MDA231_13C15NGln_shCTL_rep1-cells-polar_NMR_A protocol.id:13C15NGln_shCTL lineage_1=2_MDA231_13C15NGln_shCTL_rep1; cell_type=MDA231; plate_with_media_weight=19.66; plate_with_media_weight%units=g; replicate=1; lineage_2=2_MDA231_13C15NGln_shCTL_rep1-cells; cell_type=MDA231; protocol.id=cell_quench_collection; lineage_3=2_MDA231_13C15NGln_shCTL_rep1-cells-polar_NMR_A; protocol.id=['polar_extraction', 'NMR_file_storage2']; replicate=1; replicate%type=analytical; sample_split_ratio=0.28579888665166403; type=polar; weight=0.4825999999999999; weight%units=g; lineage_4=2_MDA231_13C15NGln_shCTL_rep1-cells-protein; dry_weight=0.0008000000000000229; dry_weight%units=g; protein_weight=0.5884; protein_weight%units=mg; protocol.id=protein_extraction; type=protein SUBJECT_SAMPLE_FACTORS - 3_MDA231_13C15NGln_shCTL_rep2-cells-polar_NMR_A protocol.id:13C15NGln_shCTL lineage_1=3_MDA231_13C15NGln_shCTL_rep2; cell_type=MDA231; plate_with_media_weight=19.65; plate_with_media_weight%units=g; replicate=2; lineage_2=3_MDA231_13C15NGln_shCTL_rep2-cells; cell_type=MDA231; protocol.id=cell_quench_collection; lineage_3=3_MDA231_13C15NGln_shCTL_rep2-cells-polar_NMR_A; protocol.id=['polar_extraction', 'NMR_file_storage3']; replicate=1; replicate%type=analytical; sample_split_ratio=0.25640876252621786; type=polar; weight=0.44010000000000016; weight%units=g; lineage_4=3_MDA231_13C15NGln_shCTL_rep2-cells-protein; dry_weight=0.0006999999999999229; dry_weight%units=g; protein_weight=0.5808; protein_weight%units=mg; protocol.id=protein_extraction; type=protein SUBJECT_SAMPLE_FACTORS - 4_MDA231_13C15NGln_shCTL_rep3-cells-polar_NMR_A protocol.id:13C15NGln_shCTL lineage_1=4_MDA231_13C15NGln_shCTL_rep3; cell_type=MDA231; plate_with_media_weight=19.17; plate_with_media_weight%units=g; replicate=3; lineage_2=4_MDA231_13C15NGln_shCTL_rep3-cells; cell_type=MDA231; protocol.id=cell_quench_collection; lineage_3=4_MDA231_13C15NGln_shCTL_rep3-cells-polar_NMR_A; protocol.id=['polar_extraction', 'NMR_file_storage4']; replicate=1; replicate%type=analytical; sample_split_ratio=0.2617664493183165; type=polar; weight=0.4416000000000002; weight%units=g; lineage_4=4_MDA231_13C15NGln_shCTL_rep3-cells-protein; dry_weight=0.0007000000000000339; dry_weight%units=g; protein_weight=0.5344000000000001; protein_weight%units=mg; protocol.id=protein_extraction; type=protein SUBJECT_SAMPLE_FACTORS - 6_MDA231_13C15NGln_shP4HA1_rep1-cells-polar_NMR_A protocol.id:13C15NGln_shP4HA1 lineage_1=6_MDA231_13C15NGln_shP4HA1_rep1; cell_type=MDA231; plate_with_media_weight=19.13; plate_with_media_weight%units=g; replicate=1; lineage_2=6_MDA231_13C15NGln_shP4HA1_rep1-cells; cell_type=MDA231; protocol.id=cell_quench_collection; lineage_3=6_MDA231_13C15NGln_shP4HA1_rep1-cells-polar_NMR_A; protocol.id=['polar_extraction', 'NMR_file_storage6']; replicate=1; replicate%type=analytical; sample_split_ratio=0.2785457645697584; type=polar; weight=0.45740000000000025; weight%units=g; lineage_4=6_MDA231_13C15NGln_shP4HA1_rep1-cells-protein; dry_weight=0.0009000000000000119; dry_weight%units=g; protein_weight=0.5952000000000001; protein_weight%units=mg; protocol.id=protein_extraction; type=protein SUBJECT_SAMPLE_FACTORS - 7_MDA231_13C15NGln_shP4HA1_rep2-cells-polar_NMR_A protocol.id:13C15NGln_shP4HA1 lineage_1=7_MDA231_13C15NGln_shP4HA1_rep2; cell_type=MDA231; plate_with_media_weight=19.66; plate_with_media_weight%units=g; replicate=2; lineage_2=7_MDA231_13C15NGln_shP4HA1_rep2-cells; cell_type=MDA231; protocol.id=cell_quench_collection; lineage_3=7_MDA231_13C15NGln_shP4HA1_rep2-cells-polar_NMR_A; protocol.id=['polar_extraction', 'NMR_file_storage7']; replicate=1; replicate%type=analytical; sample_split_ratio=0.2859185984148737; type=polar; weight=0.4797999999999998; weight%units=g; lineage_4=7_MDA231_13C15NGln_shP4HA1_rep2-cells-protein; dry_weight=0.0010999999999999899; dry_weight%units=g; protein_weight=0.5852; protein_weight%units=mg; protocol.id=protein_extraction; type=protein SUBJECT_SAMPLE_FACTORS - 8_MDA231_13C15NGln_shP4HA1_rep3-cells-polar_NMR_A protocol.id:13C15NGln_shP4HA1 lineage_1=8_MDA231_13C15NGln_shP4HA1_rep3; cell_type=MDA231; plate_with_media_weight=19.3; plate_with_media_weight%units=g; replicate=3; lineage_2=8_MDA231_13C15NGln_shP4HA1_rep3-cells; cell_type=MDA231; protocol.id=cell_quench_collection; lineage_3=8_MDA231_13C15NGln_shP4HA1_rep3-cells-polar_NMR_A; protocol.id=['polar_extraction', 'NMR_file_storage8']; replicate=1; replicate%type=analytical; sample_split_ratio=0.2918280979912975; type=polar; weight=0.48960000000000004; weight%units=g; lineage_4=8_MDA231_13C15NGln_shP4HA1_rep3-cells-protein; dry_weight=0.0008999999999999009; dry_weight%units=g; protein_weight=0.6532; protein_weight%units=mg; protocol.id=protein_extraction; type=protein #COLLECTION CO:COLLECTION_SUMMARY Collection and quench of cells in culture CO:COLLECTION_PROTOCOL_ID cell_quench_collection CO:COLLECTION_PROTOCOL_FILENAME 3A_Cells_Quench_Cell_Fan_20170412.pdf CO:SAMPLE_TYPE cultured cells #TREATMENT TR:TREATMENT_SUMMARY Cells grown in unlabeled glutamine and infected with empty vector. Cells grown TR:TREATMENT_SUMMARY in unlabeled glutamine and infected with shP4HA1 lentivirus. Cells grown in TR:TREATMENT_SUMMARY labeled glutamine and infected with empty vector. Cells grown in labeled TR:TREATMENT_SUMMARY glutamine and infected with shP4HA1 lentivirus. TR:TREATMENT_PROTOCOL_ID 12C14NGln_shCTL 12C14NGln_shP4HA1 13C15NGln_shCTL 13C15NGln_shP4HA1 #SAMPLEPREP SP:SAMPLEPREP_SUMMARY Polar extraction from cells. Protein extraction and quantification. SP:SAMPLEPREP_PROTOCOL_ID polar_extraction protein_extraction SP:SAMPLEPREP_PROTOCOL_FILENAME 4B_Extract_Polar_Lipid_Prot_Fan_070417.pdf;4D_17Jun4_Fan_Prot_Quant.pdf #CHROMATOGRAPHY #ANALYSIS AN:ANALYSIS_TYPE NMR #NMR NM:INSTRUMENT_NAME Agilent 600 NM:INSTRUMENT_TYPE FT-NMR NM:NMR_EXPERIMENT_TYPE HC-HSQC NM:STANDARD_CONCENTRATION 0.5 mM NM:SPECTROMETER_FREQUENCY 600 MHz NM:NMR_PROBE cryogenic triple resonance HCN NM:NMR_SOLVENT D20 NM:NMR_TUBE_SIZE 1.7 mm NM:SHIMMING_METHOD gradient shimming NM:PULSE_SEQUENCE gHSQCAD NM:WATER_SUPPRESSION presaturation NM:CHEMICAL_SHIFT_REF_CPD DSS NM:TEMPERATURE 15 celsius NM:ACQUISITION_TIME 2 s NM:RELAXATION_DELAY 4 s NM:BASELINE_CORRECTION_METHOD Bernstein Polynomial #NMR_METABOLITE_DATA NMR_METABOLITE_DATA:UNITS Peak area normalized NMR_METABOLITE_DATA_START Samples 2_MDA231_13C15NGln_shCTL_rep1-cells-polar_NMR_A 3_MDA231_13C15NGln_shCTL_rep2-cells-polar_NMR_A 4_MDA231_13C15NGln_shCTL_rep3-cells-polar_NMR_A 6_MDA231_13C15NGln_shP4HA1_rep1-cells-polar_NMR_A 7_MDA231_13C15NGln_shP4HA1_rep2-cells-polar_NMR_A 8_MDA231_13C15NGln_shP4HA1_rep3-cells-polar_NMR_A Factors protocol.id:13C15NGln_shCTL protocol.id:13C15NGln_shCTL protocol.id:13C15NGln_shCTL protocol.id:13C15NGln_shP4HA1 protocol.id:13C15NGln_shP4HA1 protocol.id:13C15NGln_shP4HA1 Asp-3a_1 24.5423 29.7037 24.6903 30.2698 27.1926 39.0372 Asp-3a_2 31.1144 34.9874 35.4275 38.3173 18.8921 Asp-3b_1 16.0053 19.9233 17.7178 18.3136 26.2934 12.4482 Asp-3b_2 24.0222 20.4347 18.5852 21.5341 12.1033 DSS_1 3.7089 2.7599 3.5932 3.3434 2.8324 2.8002 Gln-3_1 228.3296 181.8510 245.8179 184.8079 182.7145 298.4413 Gln-3_2 201.0946 177.7722 304.1650 294.0866 Gln-4_1 94.5777 51.5843 66.8797 75.2415 67.4374 106.0323 Gln-4_2 86.4081 53.6395 92.7602 84.8903 99.0015 88.7858 Gln+Glu-2_1 231.4167 200.0727 209.9923 175.1039 205.1693 197.4518 Glu-3-a_1 814.6414 592.3812 110.2269 700.7143 681.2972 116.9320 Glu-3-a_2 28.9067 16.8148 455.6918 7.3636 302.2954 Glu-3-a_3 30.3685 381.9717 Glu-3-a_4 36.1272 Glu-3-b_1 207.4967 622.2894 177.5475 258.9825 178.9555 278.1596 Glu-3-b_2 510.6707 342.3489 360.6655 451.9234 943.6137 Glu-4_1 261.1516 231.5831 221.2428 255.6909 231.9403 328.4375 GSH+GSSG-3_1 97.7528 183.3769 115.8869 89.3400 104.2341 44.2969 GSH+GSSG-3_2 65.8716 GSH+GSSG-4_1 49.0177 35.5730 48.5555 53.6964 42.5425 42.1479 Lac-3_1 8.3287 4.6150 7.1817 8.2858 8.7615 10.2799 P-Cho A_1 10.9800 8.0427 9.5417 10.0932 9.2622 10.8910 NMR_METABOLITE_DATA_END #METABOLITES METABOLITES_START metabolite_name pubchem_id inchi_key kegg_id other_id other_id_type ri ri_type moverz_quant Asp-3a_1 Asp-3a_2 Asp-3b_1 Asp-3b_2 DSS_1 Gln-3_1 Gln-3_2 Gln-4_1 Gln-4_2 Gln+Glu-2_1 Glu-3-a_1 Glu-3-a_2 Glu-3-a_3 Glu-3-a_4 Glu-3-b_1 Glu-3-b_2 Glu-4_1 GSH+GSSG-3_1 GSH+GSSG-3_2 GSH+GSSG-4_1 Lac-3_1 P-Cho A_1 METABOLITES_END #END