#METABOLOMICS WORKBENCH jwalejko_20200109_101642 DATATRACK_ID:1892 STUDY_ID:ST001305 ANALYSIS_ID:AN002174 VERSION 1 CREATED_ON 02-08-2024 #PROJECT PR:PROJECT_TITLE Intergrated Metabolomics and Transcriptomics Suggest Global Metabolic Response PR:PROJECT_TITLE to 2-Aminoacrylate Stress in Salmonella enterica PR:PROJECT_TYPE Comparison PR:PROJECT_SUMMARY Bacterial extractions from WT and 2-iminobutanoate/2iminopropanoate deaminase PR:PROJECT_SUMMARY (RidA) KO S. Enterica lines PR:INSTITUTE University of Georgia PR:DEPARTMENT CCRC PR:LAST_NAME Walejko PR:FIRST_NAME Jacquelyn PR:ADDRESS 315 Riverbend Road, Rm 1045, Athens GA 30602 PR:EMAIL jacquelyn.walejko@duke.edu PR:PHONE 9194792304 PR:DOI http://dx.doi.org/10.21228/M85D77 #STUDY ST:STUDY_TITLE Integrated Metabolomics and Transcriptomics Suggest the Global Metabolic ST:STUDY_TITLE Response to 2-Aminoacrylate Stress in Salmonella enterica ST:STUDY_SUMMARY NMR metabolomics of bacterial extractions from WT and ST:STUDY_SUMMARY 2-iminobutanoate/2iminopropanoate deaminase (RidA) KO S. Enterica lines ST:INSTITUTE University of Georgia ST:DEPARTMENT CCRC ST:LAST_NAME Edison ST:FIRST_NAME Arthur ST:ADDRESS 315 Riverbend Road, Athens, GA 30602 ST:EMAIL aedison@uga.edu ST:PHONE 7065428156 ST:SUBMIT_DATE 2020-01-09 #SUBJECT SU:SUBJECT_TYPE Bacteria SU:SUBJECT_SPECIES Salmonella enterica SU:TAXONOMY_ID 28901 #SUBJECT_SAMPLE_FACTORS: SUBJECT(optional)[tab]SAMPLE[tab]FACTORS(NAME:VALUE pairs separated by |)[tab]Additional sample data SUBJECT_SAMPLE_FACTORS - RidA1 Strain:RidA KO RAW_FILE_NAME=3 SUBJECT_SAMPLE_FACTORS - RidA2 Strain:RidA KO RAW_FILE_NAME=4 SUBJECT_SAMPLE_FACTORS - RidA3 Strain:RidA KO RAW_FILE_NAME=5 SUBJECT_SAMPLE_FACTORS - RidA4 Strain:RidA KO RAW_FILE_NAME=6 SUBJECT_SAMPLE_FACTORS - RidA5 Strain:RidA KO RAW_FILE_NAME=9 SUBJECT_SAMPLE_FACTORS - RidA6 Strain:RidA KO RAW_FILE_NAME=10 SUBJECT_SAMPLE_FACTORS - RidA7 Strain:RidA KO RAW_FILE_NAME=12 SUBJECT_SAMPLE_FACTORS - RidA8 Strain:RidA KO RAW_FILE_NAME=16 SUBJECT_SAMPLE_FACTORS - RidA9 Strain:RidA KO RAW_FILE_NAME=17 SUBJECT_SAMPLE_FACTORS - WT1 Strain:WT RAW_FILE_NAME=1 SUBJECT_SAMPLE_FACTORS - WT2 Strain:WT RAW_FILE_NAME=2 SUBJECT_SAMPLE_FACTORS - WT3 Strain:WT RAW_FILE_NAME=7 SUBJECT_SAMPLE_FACTORS - WT4 Strain:WT RAW_FILE_NAME=8 SUBJECT_SAMPLE_FACTORS - WT5 Strain:WT RAW_FILE_NAME=11 SUBJECT_SAMPLE_FACTORS - WT6 Strain:WT RAW_FILE_NAME=13 SUBJECT_SAMPLE_FACTORS - WT7 Strain:WT RAW_FILE_NAME=14 SUBJECT_SAMPLE_FACTORS - WT8 Strain:WT RAW_FILE_NAME=15 SUBJECT_SAMPLE_FACTORS - WT9 Strain:WT RAW_FILE_NAME=18 #COLLECTION CO:COLLECTION_SUMMARY Ten biologically independent cultures each of wild-type (DM9404) and ridA mutant CO:COLLECTION_SUMMARY (DM3480) strains were grown overnight in NB medium at 37 °C and used to CO:COLLECTION_SUMMARY inoculate (1% inoculum) 250 mL minimal glucose medium in 500 mL non-baffled CO:COLLECTION_SUMMARY flasks. Flasks were randomly arranged in an Innova®44 incubator and cultures CO:COLLECTION_SUMMARY were allowed to grow 16 h shaking at 180 RPM and 37 °C. Cultures were cooled on CO:COLLECTION_SUMMARY ice 5 min and then harvested by centrifugation at 7,000 x G for 10 min at 4 °C. CO:COLLECTION_SUMMARY The supernatant was decanted, pellets were resuspended in 10 mL ddH2O and CO:COLLECTION_SUMMARY transferred to sterile 15 mL conical tubes in which they were pelleted at 7,000 CO:COLLECTION_SUMMARY x G 10 min at 4 °C. Final supernatant was decanted and pellets were frozen in CO:COLLECTION_SUMMARY liquid nitrogen and stored at -80 °C prior to cell extractions. CO:SAMPLE_TYPE Bacterial cells CO:STORAGE_CONDITIONS -80℃ #TREATMENT TR:TREATMENT_SUMMARY These samples did not undergo addition treatment. #SAMPLEPREP SP:SAMPLEPREP_SUMMARY Each frozen bacterial pellet (~1 g) was thawed on ice and homogenized 3 times. SP:SAMPLEPREP_SUMMARY For homogenization, 2.6 g of 0.1 mm zirconium beads and 6 mL 4 C MeOH/H2O SP:SAMPLEPREP_SUMMARY (80/20) solvent was added and samples were agitated 7 times (210 s total) at SP:SAMPLEPREP_SUMMARY 1600 rpm in a FastPrep 96 (MPBIO). Samples were then centrifuged at 416 g and 4 SP:SAMPLEPREP_SUMMARY C for 16 min and supernatant was transferred to a new 15 mL conical tube. SP:SAMPLEPREP_SUMMARY Homogenization was carried out a second and third time using 4 mL MeOH/H2O with SP:SAMPLEPREP_SUMMARY 4 homogenization cycles (150 s) and 2 mL MeOH/H2O with 3 homogenization cycles SP:SAMPLEPREP_SUMMARY (150 s), respectively. Pooled supernatants from each sample were concentrated to SP:SAMPLEPREP_SUMMARY dryness using a CentriVap Benchtop Vacuum Concentrator (Labconco). The extracts SP:SAMPLEPREP_SUMMARY were reconstituted in 600 L of deuterated 100 mM sodium phosphate bu er (pH 7.4) SP:SAMPLEPREP_SUMMARY containing 1 mM of the internal standard DSS (d6 SP:SAMPLEPREP_SUMMARY 4,4-dimethyl-4-silapentane-1-sulfonic acid) and vortex mixed 2 min. Each sample SP:SAMPLEPREP_SUMMARY was transferred into 5 mm SampleJet NMR tubes for NMR analysis. #CHROMATOGRAPHY #ANALYSIS AN:LABORATORY_NAME Complex Carbohydrate Research Center NMR Facility AN:ANALYSIS_TYPE NMR AN:OPERATOR_NAME Adrien Le Guennec #NMR NM:INSTRUMENT_NAME Bruker Avance III HD NM:INSTRUMENT_TYPE FT-NMR NM:NMR_EXPERIMENT_TYPE 1D-1H NM:SPECTROMETER_FREQUENCY 800 MHz #NMR_METABOLITE_DATA NMR_METABOLITE_DATA:UNITS Area under curve NMR_METABOLITE_DATA_START Samples RidA1 RidA2 RidA3 RidA4 RidA5 RidA6 RidA7 RidA8 RidA9 WT1 WT2 WT3 WT4 WT5 WT6 WT7 WT8 WT9 Factors Strain:RidA KO Strain:RidA KO Strain:RidA KO Strain:RidA KO Strain:RidA KO Strain:RidA KO Strain:RidA KO Strain:RidA KO Strain:RidA KO Strain:WT Strain:WT Strain:WT Strain:WT Strain:WT Strain:WT Strain:WT Strain:WT Strain:WT 2-Aminobutyric acid 1.0584 1.1712 1.1595 1.0960 0.9905 1.3299 1.1182 1.1430 1.1786 1.7456 0.7645 0.8580 1.7344 1.8076 0.9397 0.8712 0.8914 0.9132 2-isopropylmalic acid 0.7678 1.4187 1.5349 0.5799 1.5595 1.5746 1.6952 1.7450 0.8039 0.3563 0.2590 0.2694 0.3355 0.4028 0.3165 0.3387 0.3208 0.3017 2-oxoglutaric acid 0.0179 0.0195 0.0176 0.0205 0.1232 0.0380 0.5135 0.0223 0.0195 0.0647 0.0223 0.4088 0.0193 0.0646 0.0904 0.0240 0.0270 0.0327 Acetate 1.8481 2.0381 1.9932 1.9802 2.1387 2.2753 2.2566 2.4532 2.0882 2.2417 2.5891 2.2490 2.2684 2.2375 2.6679 2.8998 2.6049 2.7555 Adenosine monophosphate (AMP) 0.4319 0.3632 0.3641 0.3939 0.4854 0.4068 0.4292 0.4181 0.4915 0.7440 0.8031 0.4742 0.8133 0.6335 0.9117 0.6717 0.7351 0.5333 Alanine 0.5503 0.6062 0.5957 0.7053 0.8121 0.5828 0.7299 0.8767 0.6696 0.6371 0.3598 0.6637 0.7472 0.6537 0.6007 0.4255 0.5537 0.5934 CMP 0.1456 0.1382 0.1458 0.1228 0.1166 0.1579 0.1387 0.1092 0.1341 0.0160 0.0590 0.0482 0.0221 0.0253 0.0689 0.0592 0.0668 0.0455 CTP 0.2097 0.2171 0.1859 0.1562 0.1954 0.2477 0.2412 0.1602 0.1576 0.2286 0.2312 0.1879 0.1650 0.2552 0.2287 0.2001 0.2221 0.1401 Cytidine diphosphate (CDP) 0.0221 0.0221 0.0196 0.0151 0.0099 0.0265 0.0225 0.0101 0.0076 0.0146 0.0113 0.0108 0.0055 0.0163 0.0183 0.0148 0.0170 0.0062 dCMP 0.5212 0.4366 0.4400 0.4295 0.4383 0.4502 0.4336 0.3937 0.4513 0.7172 0.7915 0.6448 0.7114 0.6160 0.7455 0.6756 0.6992 0.5144 Ethanolamine 0.0286 0.0315 0.0381 0.0426 0.0472 0.0370 0.0333 0.0815 0.0401 0.0259 0.0553 0.0352 0.0632 0.0325 0.0149 0.0344 0.0320 0.0368 Formate 0.1223 0.0983 0.1143 0.1165 0.1002 0.1378 0.1189 0.1352 0.1640 0.1664 0.1759 0.1497 0.1676 0.1712 0.1356 0.1856 0.1598 0.1497 Glucose 0.0533 0.0638 0.0640 0.0604 0.0584 0.0630 0.0580 0.0491 0.0571 0.0460 0.0486 0.0437 0.0464 0.0497 0.0325 0.0380 0.0339 0.0369 Glutamate 0.2275 0.3026 0.2800 0.2176 0.2214 0.2853 0.1233 0.3434 0.2523 0.2080 0.1511 0.1117 0.2092 0.2175 0.2131 0.1588 0.1932 0.1401 GSSG 2.0290 2.1001 2.0994 1.7015 1.7569 2.2205 2.2245 2.0262 1.9906 0.7047 0.2959 0.2944 0.6457 0.9278 0.5267 0.5120 0.5076 0.4752 Isoleucine 0.2421 0.2201 0.2169 0.2532 0.2183 0.2035 0.2265 0.2207 0.2125 0.2525 0.3953 0.5277 0.2446 0.2316 0.5031 0.3541 0.4963 0.3261 Leucine 0.8115 0.8509 0.8682 0.8873 0.7454 0.8407 0.7513 0.7440 0.7443 1.9082 1.8209 2.3367 1.7510 1.8290 2.1424 1.6953 2.1054 1.5501 Lysine 0.0760 0.0860 0.0869 0.0784 0.0776 0.0445 0.0436 0.0661 0.0524 0.5182 0.7595 0.4593 0.5702 0.4370 0.7738 0.6007 0.6878 0.4919 Malic acid 0.0287 0.0283 0.0279 0.0354 0.0370 0.0324 0.0359 0.0309 0.0299 0.0315 0.0404 0.0432 0.0347 0.0339 0.0278 0.0397 0.0331 0.0424 Maltose 1.4993 1.5102 1.5347 1.6495 1.6806 1.5195 1.7021 1.7472 1.5210 1.3653 1.4268 1.4939 1.2983 1.3343 1.6250 1.4356 1.4861 1.2618 Methionine 0.1694 0.1544 0.1693 0.1620 0.1510 0.1670 0.1558 0.1855 0.1831 0.0767 0.0673 0.0596 0.0808 0.0923 0.0754 0.0779 0.0750 0.0752 N-acetyl alanine 0.0254 0.0202 0.0190 0.0261 0.0210 0.0216 0.0261 0.0181 0.0163 0.0103 0.0176 0.0261 0.0112 0.0146 0.0217 0.0192 0.0217 0.0143 N-Acetylputrescine 0.1241 0.1313 0.1258 0.1209 0.1026 0.1281 0.0978 0.1182 0.1546 0.1645 0.1854 0.1067 0.1726 0.1565 0.1562 0.1765 0.1521 0.1650 Nicotinamide Adenine Dinucleotide (NAD) 0.0807 0.0867 0.0846 0.0560 0.0541 0.1362 0.1047 0.0544 0.0642 0.1171 0.0941 0.0436 0.0486 0.1208 0.1201 0.0881 0.1171 0.0358 Nicotinamide Adenine Dinucleotide Phosphate (NADP) 0.0430 0.0354 0.0381 0.0494 0.0383 0.0392 0.0392 0.0339 0.0363 0.0595 0.0541 0.0611 0.0533 0.0563 0.0431 0.0534 0.0503 0.0431 Nicotinate Mononucleotide 0.3938 0.3597 0.3709 0.4705 0.4138 0.3277 0.3934 0.2712 0.2546 0.3729 0.2068 0.4592 0.3230 0.3743 0.2796 0.3419 0.2931 0.4423 Pantothenic acid 0.1765 0.1438 0.1429 0.1708 0.1391 0.1461 0.1397 0.1683 0.1888 0.1095 0.1452 0.1499 0.1195 0.1137 0.1697 0.1295 0.1532 0.1190 Phenylalanine 0.0157 0.0184 0.0164 0.0192 0.0182 0.0125 0.0127 0.0149 0.0103 0.1170 0.1216 0.1772 0.1072 0.1074 0.1725 0.1061 0.1647 0.0930 Putrescine 0.2158 0.4245 0.3547 0.1901 0.2263 0.3028 0.2017 0.5358 0.5922 0.4056 0.6203 0.2397 0.4427 0.2501 0.9181 0.4287 0.5458 0.3682 Serine 0.0627 0.0537 0.0543 0.0454 0.0596 0.0518 0.0583 0.0447 0.0498 0.0654 0.0610 0.0611 0.0648 0.0707 0.0511 0.0625 0.0558 0.0583 Succinic acid 1.4985 1.6002 1.6206 1.5791 1.8176 1.7450 1.9511 1.8998 1.3421 2.6761 1.8942 2.0507 2.5313 2.5102 3.0336 2.5934 2.7424 2.3923 Threonine 0.0716 0.0699 0.0708 0.0631 0.0536 0.0809 0.0711 0.0494 0.0507 0.0527 0.0558 0.0480 0.0455 0.0559 0.0629 0.0555 0.0598 0.0383 Tyrosine 0.0398 0.0432 0.0400 0.0377 0.0427 0.0490 0.0502 0.0467 0.0477 0.0367 0.0427 0.0701 0.0372 0.0434 0.0105 0.0455 0.0493 0.0399 UMP 0.1592 0.1776 0.1780 0.1648 0.1916 0.1440 0.1453 0.2037 0.1916 0.1532 0.2560 0.2086 0.1917 0.1335 0.1643 0.2105 0.1715 0.1853 Uracil 0.0483 0.0477 0.0486 0.0751 0.1032 0.0421 0.0721 0.1315 0.0943 0.0588 0.0460 0.0711 0.0732 0.0584 0.0429 0.0465 0.0464 0.0866 Uridine 5^-triphosphate (UTP) 0.0661 0.0600 0.0566 0.0549 0.0575 0.0760 0.0802 0.0451 0.0467 0.0380 0.0330 0.0427 0.0195 0.0506 0.0610 0.0468 0.0589 0.0299 Valine 2.7151 2.5415 2.4652 2.8657 2.6754 2.7362 2.7444 2.8655 2.7751 1.0776 1.1239 1.2511 1.0727 1.2231 1.4669 1.2963 1.3788 1.2067 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 2-Aminobutyric acid C02356 2-isopropylmalic acid C02504 2-oxoglutaric acid C00026 Acetate C00033 Adenosine monophosphate (AMP) C00020 Alanine C00041 CMP C00055 CTP C00063 Cytidine diphosphate (CDP) C00112 dCMP C00239 Ethanolamine C00189 Formate C00058 Glucose C00031 Glutamate C00025 GSSG C00127 Isoleucine C00407 Leucine C00123 Lysine C00047 Malic acid C00711 Maltose C00208 Methionine C00073 N-acetyl alanine C04341 N-Acetylputrescine C02714 Nicotinamide Adenine Dinucleotide (NAD) C00003 Nicotinamide Adenine Dinucleotide Phosphate (NADP) C00006 Nicotinate Mononucleotide C01185 Pantothenic acid C00864 Phenylalanine C00079 Putrescine C00134 Serine C00065 Succinic acid C00042 Threonine C00188 Tyrosine C00082 UMP C00105 Uracil C00106 Uridine 5'-triphosphate (UTP) C00075 Valine C00183 METABOLITES_END #END