#METABOLOMICS WORKBENCH KasparasP_20240820_021723 DATATRACK_ID:5125 STUDY_ID:ST003418 ANALYSIS_ID:AN005617 PROJECT_ID:PR002115 VERSION 1 CREATED_ON August 20, 2024, 4:02 am #PROJECT PR:PROJECT_TITLE TRAM–LAG1–CLN8 family proteins are acyltransferases regulating phospholipid PR:PROJECT_TITLE composition PR:PROJECT_SUMMARY The diversity of cellular phospholipids, crucial for membrane homeostasis and PR:PROJECT_SUMMARY function, arises from enzymatic remodeling of their fatty acyl chains. In this PR:PROJECT_SUMMARY work, we reveal that poorly understood TRAM–LAG1–CLN8 domain-containing PR:PROJECT_SUMMARY (TLCD) proteins are phospholipid remodeling enzymes. We demonstrate that TLCD1 PR:PROJECT_SUMMARY is an evolutionarily conserved lysophosphatidylethanolamine acyltransferase, PR:PROJECT_SUMMARY which regulates cellular phospholipid composition and generates novel fatty acid PR:PROJECT_SUMMARY and thiamine (vitamin B1) esters as its secondary products. Furthermore, we PR:PROJECT_SUMMARY establish that human TLCD protein CLN8, mutations in which cause fatal PR:PROJECT_SUMMARY neurodegenerative Batten disease, is a lysophosphatidylglycerol acyltransferase. PR:PROJECT_SUMMARY We show that CLN8 catalyzes the essential step in the biosynthesis of PR:PROJECT_SUMMARY bis(monoacylglycero)phosphate, a phospholipid critical for lysosome function. PR:PROJECT_SUMMARY Our study unveils a new family of acyltransferases integral to cellular membrane PR:PROJECT_SUMMARY phospholipid homeostasis and human disease. PR:INSTITUTE University of Cambridge PR:LAST_NAME Petkevicius PR:FIRST_NAME Kasparas PR:ADDRESS The Keith Peters Building, Cambridge, Cambridgeshire, CB2 0XY, United Kingdom PR:EMAIL kp416@mrc-mbu.cam.ac.uk PR:PHONE +447500233355 PR:PUBLICATIONS TRAM–LAG1–CLN8 family proteins are acyltransferases regulating phospholipid PR:PUBLICATIONS composition #STUDY ST:STUDY_TITLE Analysis of lipid composition of TLCD1 KO cultured cells ST:STUDY_SUMMARY Untargeted lipidomics experiments were conducted to analyze the differences in ST:STUDY_SUMMARY lipid composition between TLCD1 KO and isogenic control HeLa and U2OS cell ST:STUDY_SUMMARY lines. See a reference publication (Sheokand et al) for details. ST:INSTITUTE University of Cambridge ST:LAST_NAME Petkevicius ST:FIRST_NAME Kasparas ST:ADDRESS The Keith Peters Building ST:EMAIL kp416@mrc-mbu.cam.ac.uk ST:PHONE 07500233355 #SUBJECT SU:SUBJECT_TYPE Cultured cells SU:SUBJECT_SPECIES Homo sapiens SU:TAXONOMY_ID 9606 #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 - Cell process blank Genotype:Process blank | Sample source:Process blank | Clonality:Process blank | Replicate:Process blank RAW_FILE_NAME(Raw file name)=Cell_process_blank.raw SUBJECT_SAMPLE_FACTORS - HeLa Wild-Type poolReplicate 1 Genotype:Wild-type | Sample source:HeLa | Clonality:Cell pool | Replicate:1 RAW_FILE_NAME(Raw file name)=HeLa_Wild-Type_pool_Replicate_1.raw SUBJECT_SAMPLE_FACTORS - HeLa Wild-Type poolReplicate 2 Genotype:Wild-type | Sample source:HeLa | Clonality:Cell pool | Replicate:2 RAW_FILE_NAME(Raw file name)=HeLa_Wild-Type_pool_Replicate_2.raw SUBJECT_SAMPLE_FACTORS - HeLa WildType poolReplicate 3 Genotype:Wild-type | Sample source:HeLa | Clonality:Cell pool | Replicate:3 RAW_FILE_NAME(Raw file name)=HeLa_Wild-Type_pool_Replicate_3.raw SUBJECT_SAMPLE_FACTORS - HeLa TLCD1 KO poolReplicate 1 Genotype:TLCD1 KO | Sample source:HeLa | Clonality:Cell pool | Replicate:1 RAW_FILE_NAME(Raw file name)=HeLa_TLCD1_KO_pool_Replicate_1.raw SUBJECT_SAMPLE_FACTORS - HeLa TLCD1 KO poolReplicate 2 Genotype:TLCD1 KO | Sample source:HeLa | Clonality:Cell pool | Replicate:2 RAW_FILE_NAME(Raw file name)=HeLa_TLCD1_KO_pool_Replicate_2.raw SUBJECT_SAMPLE_FACTORS - HeLa TLCD1 KO poolReplicate 3 Genotype:TLCD1 KO | Sample source:HeLa | Clonality:Cell pool | Replicate:3 RAW_FILE_NAME(Raw file name)=HeLa_TLCD1_KO_pool_Replicate_3.raw SUBJECT_SAMPLE_FACTORS - HeLa WildType clone 1Replicate 1 Genotype:Wild-type | Sample source:HeLa | Clonality:Clone derived | Replicate:1 RAW_FILE_NAME(Raw file name)=HeLa_Wild-Type_clone_1_Replicate_1.raw SUBJECT_SAMPLE_FACTORS - HeLa Wild-Type clone 1Replicate 2 Genotype:Wild-type | Sample source:HeLa | Clonality:Clone derived | Replicate:2 RAW_FILE_NAME(Raw file name)=HeLa_Wild-Type_clone_1_Replicate_2.raw SUBJECT_SAMPLE_FACTORS - HeLa Wild-Type clone 1Replicate 3 Genotype:Wild-type | Sample source:HeLa | Clonality:Clone derived | Replicate:3 RAW_FILE_NAME(Raw file name)=HeLa_Wild-Type_clone_1_Replicate_3.raw SUBJECT_SAMPLE_FACTORS - HeLa Wild-Type clone 2Replicate 1 Genotype:Wild-type | Sample source:HeLa | Clonality:Clone derived | Replicate:1 RAW_FILE_NAME(Raw file name)=HeLa_Wild-Type_clone_2_Replicate_1.raw SUBJECT_SAMPLE_FACTORS - HeLa WildType clone 2Replicate 2 Genotype:Wild-type | Sample source:HeLa | Clonality:Clone derived | Replicate:2 RAW_FILE_NAME(Raw file name)=HeLa_Wild-Type_clone_2_Replicate_2.raw SUBJECT_SAMPLE_FACTORS - HeLa Wild-Type clone 2Replicate 3 Genotype:Wild-type | Sample source:HeLa | Clonality:Clone derived | Replicate:3 RAW_FILE_NAME(Raw file name)=HeLa_Wild-Type_clone_2_Replicate_3.raw SUBJECT_SAMPLE_FACTORS - HeLa TLCD1 KO cloneReplicate 1 Genotype:TLCD1 KO | Sample source:HeLa | Clonality:Clone derived | Replicate:1 RAW_FILE_NAME(Raw file name)=HeLa_TLCD1_KO_clone_Replicate_1.raw SUBJECT_SAMPLE_FACTORS - HeLa TLCD1 KO cloneReplicate 2 Genotype:TLCD1 KO | Sample source:HeLa | Clonality:Clone derived | Replicate:2 RAW_FILE_NAME(Raw file name)=HeLa_TLCD1_KO_clone_Replicate_2.raw SUBJECT_SAMPLE_FACTORS - HeLa TLCD1 KO cloneReplicate 3 Genotype:TLCD1 KO | Sample source:HeLa | Clonality:Clone derived | Replicate:3 RAW_FILE_NAME(Raw file name)=HeLa_TLCD1_KO_clone_Replicate_3.raw SUBJECT_SAMPLE_FACTORS - U20S WildType poolReplicate 1 Genotype:Wild-type | Sample source:U2OS | Clonality:Cell pool | Replicate:1 RAW_FILE_NAME(Raw file name)=U20S_Wild-Type_pool_Replicate_1.raw SUBJECT_SAMPLE_FACTORS - U20S Wild-Type poolReplicate 2 Genotype:Wild-type | Sample source:U2OS | Clonality:Cell pool | Replicate:2 RAW_FILE_NAME(Raw file name)=U20S_Wild-Type_pool_Replicate_2.raw SUBJECT_SAMPLE_FACTORS - U20S Wild-Type poolReplicate 3 Genotype:Wild-type | Sample source:U2OS | Clonality:Cell pool | Replicate:3 RAW_FILE_NAME(Raw file name)=U20S_Wild-Type_pool_Replicate_3.raw SUBJECT_SAMPLE_FACTORS - U20S TLCD1 KO poolReplicate 1 Genotype:TLCD1 KO | Sample source:U2OS | Clonality:Cell pool | Replicate:1 RAW_FILE_NAME(Raw file name)=U20S_TLCD1_KO_pool_Replicate_1.raw SUBJECT_SAMPLE_FACTORS - U20S TLCD1 KO poolReplicate 2 Genotype:TLCD1 KO | Sample source:U2OS | Clonality:Cell pool | Replicate:2 RAW_FILE_NAME(Raw file name)=U20S_TLCD1_KO_pool_Replicate_2.raw SUBJECT_SAMPLE_FACTORS - U20S TLCD1 KO poolReplicate 3 Genotype:TLCD1 KO | Sample source:U2OS | Clonality:Cell pool | Replicate:3 RAW_FILE_NAME(Raw file name)=U20S_TLCD1_KO_pool_Replicate_3.raw SUBJECT_SAMPLE_FACTORS - U20S Wild-Type clone 1Replicate 1 Genotype:Wild-type | Sample source:U2OS | Clonality:Clone derived | Replicate:1 RAW_FILE_NAME(Raw file name)=U20S_Wild-Type_clone_1_Replicate_1.raw SUBJECT_SAMPLE_FACTORS - U20S Wild-Type clone 1Replicate 2 Genotype:Wild-type | Sample source:U2OS | Clonality:Clone derived | Replicate:2 RAW_FILE_NAME(Raw file name)=U20S_Wild-Type_clone_1_Replicate_2.raw SUBJECT_SAMPLE_FACTORS - U20S Wild-Type clone 1Replicate 3 Genotype:Wild-type | Sample source:U2OS | Clonality:Clone derived | Replicate:3 RAW_FILE_NAME(Raw file name)=U20S_Wild-Type_clone_1_Replicate_3.raw SUBJECT_SAMPLE_FACTORS - U20S Wild-Type clone 2Replicate 1 Genotype:Wild-type | Sample source:U2OS | Clonality:Clone derived | Replicate:1 RAW_FILE_NAME(Raw file name)=U20S_Wild-Type_clone_2_Replicate_1.raw SUBJECT_SAMPLE_FACTORS - U20S Wild-Type clone 2Replicate 2 Genotype:Wild-type | Sample source:U2OS | Clonality:Clone derived | Replicate:2 RAW_FILE_NAME(Raw file name)=U20S_Wild-Type_clone_2_Replicate_2.raw SUBJECT_SAMPLE_FACTORS - U20S Wild-Type clone 2Replicate 3 Genotype:Wild-type | Sample source:U2OS | Clonality:Clone derived | Replicate:3 RAW_FILE_NAME(Raw file name)=U20S_Wild-Type_clone_2_Replicate_3.raw SUBJECT_SAMPLE_FACTORS - U20S TLCD1 KO cloneReplicate 1 Genotype:TLCD1 KO | Sample source:U2OS | Clonality:Clone derived | Replicate:1 RAW_FILE_NAME(Raw file name)=U20S_TLCD1_KO_clone_Replicate_1.raw SUBJECT_SAMPLE_FACTORS - U20S TLCD1 KO cloneReplicate 2 Genotype:TLCD1 KO | Sample source:U2OS | Clonality:Clone derived | Replicate:2 RAW_FILE_NAME(Raw file name)=U20S_TLCD1_KO_clone_Replicate_2.raw SUBJECT_SAMPLE_FACTORS - U20S TLCD1 KO cloneReplicate 3 Genotype:TLCD1 KO | Sample source:U2OS | Clonality:Clone derived | Replicate:3 RAW_FILE_NAME(Raw file name)=U20S_TLCD1_KO_clone_Replicate_3.raw #COLLECTION CO:COLLECTION_SUMMARY All cell lines were maintained in Dulbecco’s Modified Eagle Medium (DMEM, CO:COLLECTION_SUMMARY 11965084, Thermo Fisher) supplemented with 10% fetal bovine serum (FBS, Sigma) CO:COLLECTION_SUMMARY at 37°C and 5% CO2. To collect samples for lipid measurements, confluent cells CO:COLLECTION_SUMMARY in 6-well plates were swiftly washed twice with 3 ml of ice-cold PBS on ice, CO:COLLECTION_SUMMARY then scraped into 500 μl of ice-cold PBS on ice using inverted 200 μl pipette CO:COLLECTION_SUMMARY tip. The resulting cell suspension was transferred into pre-chilled 2 ml plastic CO:COLLECTION_SUMMARY vials used for lipid extraction (3469-11, Thermo Scientific), and centrifuged at CO:COLLECTION_SUMMARY 500 g, 4°C for 5 min. The supernatant was aspirated and the cell pellet was CO:COLLECTION_SUMMARY snap-frozen and stored at -70°C. One well of a 6-well plate yielded one CO:COLLECTION_SUMMARY replicate in analysis. CO:SAMPLE_TYPE Cultured cells CO:STORAGE_CONDITIONS -80℃ #TREATMENT TR:TREATMENT_SUMMARY Cells were untreated (collected under standard culturing conditions). #SAMPLEPREP SP:SAMPLEPREP_SUMMARY The extraction of total lipids from cellular matrices, immunoprecipitated SP:SAMPLEPREP_SUMMARY mitochondria and in vitro assays was conducted employing the butanol-methanol SP:SAMPLEPREP_SUMMARY (BUME) method, as described in detail (37). Note that the chloroform-methanol SP:SAMPLEPREP_SUMMARY lipid extraction methods result in the partitioning of acyl-thiamines into the SP:SAMPLEPREP_SUMMARY polar phase. 2 ml screw cap plastic tubes were used (3469-11, Thermo SP:SAMPLEPREP_SUMMARY Scientific), and a blank extraction was always performed in parallel to account SP:SAMPLEPREP_SUMMARY for the plastic-related contaminants. The extraction commenced with the SP:SAMPLEPREP_SUMMARY homogenization of frozen cell pellets, mitochondrial beads or in vitro assay SP:SAMPLEPREP_SUMMARY mixtures in a 0.5 ml ice-cold solution of butanol to methanol in a 3:1 ratio. SP:SAMPLEPREP_SUMMARY For lipidomics samples, BUME solution was enriched with SPLASH internal standard SP:SAMPLEPREP_SUMMARY mix (330707, Avanti Polar Lipids). For the extraction, a further 0.5 ml of 1% SP:SAMPLEPREP_SUMMARY acetic acid and 0.5 ml of a heptane:ethyl acetate 3:1 mixture were added, SP:SAMPLEPREP_SUMMARY followed by vigorous vortexing for a total of 5 minutes. The mixture was then SP:SAMPLEPREP_SUMMARY centrifuged at 6000 g for 5 minutes, allowing for the separation of phases, SP:SAMPLEPREP_SUMMARY after which the upper organic phase was carefully decanted into glass vials. A SP:SAMPLEPREP_SUMMARY second extraction was conducted on the remaining aqueous phase, with the newly SP:SAMPLEPREP_SUMMARY acquired upper phase being combined in the same glass vials as the first. Post SP:SAMPLEPREP_SUMMARY extraction, the solvents were evaporated under a stream of nitrogen, and the SP:SAMPLEPREP_SUMMARY resultant dry lipid extracts were preserved at −70°C pending further SP:SAMPLEPREP_SUMMARY analysis. #CHROMATOGRAPHY CH:CHROMATOGRAPHY_SUMMARY Dried samples were reconstituted in 100 μL of isopropanol, acetonitrile, and CH:CHROMATOGRAPHY_SUMMARY water (2:1:1 ratio) and thoroughly vortexed. Liquid chromatography was conducted CH:CHROMATOGRAPHY_SUMMARY using a Shimadzu HPLC System, with 10 μL of the sample introduced onto a Waters CH:CHROMATOGRAPHY_SUMMARY Acquity Premier UPLC® CSH column (1.7 μm pore size, 2.1 mm × 50 mm), which CH:CHROMATOGRAPHY_SUMMARY was maintained at 55 °C. The mobile phase A comprised a 6:4 ratio of CH:CHROMATOGRAPHY_SUMMARY acetonitrile to water with 10 mM ammonium formate, and mobile phase B consisted CH:CHROMATOGRAPHY_SUMMARY of a 9:1 ratio of isopropanol to acetonitrile with 10 mM ammonium formate. A CH:CHROMATOGRAPHY_SUMMARY flow rate of 500 μL per minute was maintained with a gradient protocol for CH:CHROMATOGRAPHY_SUMMARY mobile phase B as follows: 0.00 minutes_40% mobile phase B; 1.5 minutes_40% CH:CHROMATOGRAPHY_SUMMARY mobile phase B; 8.00 minutes_99% mobile phase B; 10.00 minutes_99% mobile phase CH:CHROMATOGRAPHY_SUMMARY B; 10.10 minutes_40% mobile phase B; 12.00 minutes_40% mobile phase. The sample CH:CHROMATOGRAPHY_SUMMARY injection needle was rinsed with a 9:1 isopropanol and acetonitrile solution CH:CHROMATOGRAPHY_SUMMARY (strong wash) and isopropanol, acetonitrile, and water (2:1:1, weak wash). CH:CHROMATOGRAPHY_TYPE Reversed phase CH:INSTRUMENT_NAME Waters Acquity CH:COLUMN_NAME Waters ACQUITY UPLC CSH C18 (100 x 2.1mm,1.7um) CH:SOLVENT_A 6:4 of acetonitrile to water with 10 mM ammonium formate CH:SOLVENT_B 9:1 of isopropanol to acetonitrile with 10 mM ammonium formate CH:FLOW_GRADIENT Gradient protocol for mobile phase B as follows: 0.00 minutes_40% mobile phase CH:FLOW_GRADIENT B; 1.5 minutes_40% mobile phase B; 8.00 minutes_99% mobile phase B; 10.00 CH:FLOW_GRADIENT minutes_99% mobile phase B; 10.10 minutes_40% mobile phase B; 12.00 minutes_40% CH:FLOW_GRADIENT mobile phase. CH:FLOW_RATE 500 μL per minute CH:COLUMN_TEMPERATURE 55 #ANALYSIS AN:ANALYSIS_TYPE MS #MS MS:INSTRUMENT_NAME Thermo Q Exactive Orbitrap MS:INSTRUMENT_TYPE Orbitrap MS:MS_TYPE ESI MS:ION_MODE UNSPECIFIED MS:MS_COMMENTS Mass spectrometric detection was carried out on a ThermoFisher Scientific MS:MS_COMMENTS Q-Exactive Orbitrap equipped with a heated electrospray ionization source. The MS:MS_COMMENTS mass spectrometer was calibrated immediately before sample analysis using MS:MS_COMMENTS positive and negative ionization calibration solution (recommended by Thermo MS:MS_COMMENTS Scientific). The electrospray ionization parameters were optimized with a 50:50 MS:MS_COMMENTS mix of mobile phase A and B for spray stability, setting the capillary MS:MS_COMMENTS temperature at 300 °C, source heater temperature at 420 °C, with the sheath, MS:MS_COMMENTS auxiliary, and spare gas flows at specific arbitrary units (40, 15 and 3, MS:MS_COMMENTS respectively), and source voltage at 4 kV. The mass spectrometer operated at a MS:MS_COMMENTS scan rate of 4 Hz, yielding a resolution of 35,000 at m/z 200, over a full-scan MS:MS_COMMENTS range from m/z 120 to 1800, with continuous switching between positive and MS:MS_COMMENTS negative modes. #MS_METABOLITE_DATA MS_METABOLITE_DATA:UNITS nmol MS_METABOLITE_DATA_START Samples Cell process blank HeLa Wild-Type poolReplicate 1 HeLa Wild-Type poolReplicate 2 HeLa WildType poolReplicate 3 HeLa TLCD1 KO poolReplicate 1 HeLa TLCD1 KO poolReplicate 2 HeLa TLCD1 KO poolReplicate 3 HeLa WildType clone 1Replicate 1 HeLa Wild-Type clone 1Replicate 2 HeLa Wild-Type clone 1Replicate 3 HeLa Wild-Type clone 2Replicate 1 HeLa WildType clone 2Replicate 2 HeLa Wild-Type clone 2Replicate 3 HeLa TLCD1 KO cloneReplicate 1 HeLa TLCD1 KO cloneReplicate 2 HeLa TLCD1 KO cloneReplicate 3 U20S WildType poolReplicate 1 U20S Wild-Type poolReplicate 2 U20S Wild-Type poolReplicate 3 U20S TLCD1 KO poolReplicate 1 U20S TLCD1 KO poolReplicate 2 U20S TLCD1 KO poolReplicate 3 U20S Wild-Type clone 1Replicate 1 U20S Wild-Type clone 1Replicate 2 U20S Wild-Type clone 1Replicate 3 U20S Wild-Type clone 2Replicate 1 U20S Wild-Type clone 2Replicate 2 U20S Wild-Type clone 2Replicate 3 U20S TLCD1 KO cloneReplicate 1 U20S TLCD1 KO cloneReplicate 2 U20S TLCD1 KO cloneReplicate 3 Factors Genotype:Process blank | Sample source:Process blank | Clonality:Process blank | Replicate:Process blank Genotype:Wild-type | Sample source:HeLa | Clonality:Cell pool | Replicate:1 Genotype:Wild-type | Sample source:HeLa | Clonality:Cell pool | Replicate:2 Genotype:Wild-type | Sample source:HeLa | Clonality:Cell pool | Replicate:3 Genotype:TLCD1 KO | Sample source:HeLa | Clonality:Cell pool | Replicate:1 Genotype:TLCD1 KO | Sample source:HeLa | Clonality:Cell pool | Replicate:2 Genotype:TLCD1 KO | Sample source:HeLa | Clonality:Cell pool | Replicate:3 Genotype:Wild-type | Sample source:HeLa | Clonality:Clone derived | Replicate:1 Genotype:Wild-type | Sample source:HeLa | Clonality:Clone derived | Replicate:2 Genotype:Wild-type | Sample source:HeLa | Clonality:Clone derived | Replicate:3 Genotype:Wild-type | Sample source:HeLa | Clonality:Clone derived | Replicate:1 Genotype:Wild-type | Sample source:HeLa | Clonality:Clone derived | Replicate:2 Genotype:Wild-type | Sample source:HeLa | Clonality:Clone derived | Replicate:3 Genotype:TLCD1 KO | Sample source:HeLa | Clonality:Clone derived | Replicate:1 Genotype:TLCD1 KO | Sample source:HeLa | Clonality:Clone derived | Replicate:2 Genotype:TLCD1 KO | Sample source:HeLa | Clonality:Clone derived | Replicate:3 Genotype:Wild-type | Sample source:U2OS | Clonality:Cell pool | Replicate:1 Genotype:Wild-type | Sample source:U2OS | Clonality:Cell pool | Replicate:2 Genotype:Wild-type | Sample source:U2OS | Clonality:Cell pool | Replicate:3 Genotype:TLCD1 KO | Sample source:U2OS | Clonality:Cell pool | Replicate:1 Genotype:TLCD1 KO | Sample source:U2OS | Clonality:Cell pool | Replicate:2 Genotype:TLCD1 KO | Sample source:U2OS | Clonality:Cell pool | Replicate:3 Genotype:Wild-type | Sample source:U2OS | Clonality:Clone derived | Replicate:1 Genotype:Wild-type | Sample source:U2OS | Clonality:Clone derived | Replicate:2 Genotype:Wild-type | Sample source:U2OS | Clonality:Clone derived | Replicate:3 Genotype:Wild-type | Sample source:U2OS | Clonality:Clone derived | Replicate:1 Genotype:Wild-type | Sample source:U2OS | Clonality:Clone derived | Replicate:2 Genotype:Wild-type | Sample source:U2OS | Clonality:Clone derived | Replicate:3 Genotype:TLCD1 KO | Sample source:U2OS | Clonality:Clone derived | Replicate:1 Genotype:TLCD1 KO | Sample source:U2OS | Clonality:Clone derived | Replicate:2 Genotype:TLCD1 KO | Sample source:U2OS | Clonality:Clone derived | Replicate:3 PE_(34:1) 1022823 0.187901734 0.145045452 0.201884295 0.121710297 0.148609138 0.163156933 0.227184022 0.215303936 0.224973072 0.220915227 0.284269637 0.277423322 0.101066772 0.085050763 0.08621723 0.249174314 0.171312194 0.217500263 0.2425762 0.293209339 0.28081422 0.195599689 0.192477413 0.16861707 0.209711903 0.180160925 0.224548606 0.145048647 0.211907038 0.194254151 PE_(34:2) 0 0.018576298 0.021360159 0.029232169 0.007429292 0.009259734 0.008344382 0.033624231 0.034483564 0.03479247 0.04161393 0.047953093 0.042160136 0.00196992 0.001721508 0.001560944 0.043738298 0.034853305 0.041869307 0.022901036 0.027021184 0.021983773 0.027696697 0.031449308 0.026062347 0.032907673 0.02940544 0.034139336 0.009068567 0.005561351 0.015383424 PE_(35:1) 272498 0.03392432 0.030257486 0.036468866 0.027121343 0.032579222 0.033892533 0.047710614 0.046697203 0.04752163 0.038514969 0.041126654 0.036866298 0.02759117 0.024937862 0.023933166 0.037023057 0.024234244 0.032710238 0.039920641 0.04301159 0.041281647 0.037610453 0.03731011 0.033115758 0.031860189 0.024684524 0.030116346 0.035252438 0.047438814 0.04492636 PE_(36:1) 1536460 0.381565058 0.322105288 0.408133661 0.332517851 0.367599103 0.440286673 0.54839163 0.496553575 0.51432126 0.444121541 0.426000296 0.428511383 0.397191549 0.330619561 0.335062206 0.306136755 0.193853174 0.25475438 0.370095943 0.399737786 0.457465543 0.326154728 0.321833041 0.290190295 0.324060585 0.23672395 0.30730953 0.27546516 0.39113438 0.347736357 PE_(36:2) 650733 0.254421712 0.229013588 0.265373222 0.146610845 0.161508418 0.164420109 0.32622425 0.328835838 0.316917438 0.306245994 0.328014362 0.321166626 0.081594233 0.060050969 0.061846993 0.277856931 0.204486278 0.246090334 0.190957975 0.209655371 0.207343296 0.275244115 0.272814421 0.249452773 0.223300489 0.192612162 0.234431927 0.100841801 0.143412525 0.198162178 PE_(36:3) 0 0.013242189 0.011386534 0.008631244 0.006520997 0.006705991 0.007991754 0.013540787 0.009073252 0.007521008 0.011913573 0.012840785 0.011234469 0.001430165 0.001737679 0.001379733 0.029588736 0.018813537 0.023070661 0.021755772 0.023630543 0.024187214 0.02056614 0.024223742 0.019336888 0.023953012 0.018891364 0.020970933 0.013471995 0.019160092 0.01941333 PE_(36:4) 108448 0.098675606 0.086852834 0.115249431 0.084316503 0.088692033 0.09617645 0.110089835 0.101573367 0.106655852 0.103158977 0.118147713 0.101715319 0.061266149 0.050209852 0.046297808 0.110139128 0.089181207 0.100620073 0.12014884 0.135639644 0.130485071 0.116148093 0.116743121 0.096053667 0.105333842 0.091215439 0.107616636 0.078706092 0.131025732 0.124401807 PE_(36:5) 0 0.029752244 0.025020508 0.034580699 0.009459883 0.010408392 0.013476884 0.03548983 0.034811272 0.037560496 0.032416618 0.0327764 0.034477074 0.005581015 0.005285744 0.004588553 0.043267488 0.030461982 0.039286878 0.016675715 0.018396025 0.020012868 0.037933622 0.038957826 0.035714486 0.03819488 0.034162421 0.041593013 0.005306083 0.006863041 0.004926828 PE_(38:3) 577818 0.051907505 0.04445581 0.054800028 0.041070697 0.047389536 0.049145993 0.060591444 0.062682934 0.057191951 0.060800089 0.066205966 0.056907762 0.023202463 0.021858708 0.019876429 0.085985494 0.059715753 0.078316595 0.083615072 0.092889335 0.085879602 0.089852994 0.087791743 0.078091995 0.100952303 0.080055813 0.092160441 0.088333159 0.11187958 0.108737619 PE_(38:4) 3200829 0.434175245 0.377441019 0.443370263 0.446195272 0.474359652 0.518613302 0.456643278 0.458667297 0.456517782 0.353360167 0.365351587 0.352110601 0.429985037 0.377600625 0.343058031 0.471473247 0.353944751 0.419104152 0.570428964 0.610278342 0.611553291 0.519131714 0.523134099 0.474948594 0.468232247 0.422827422 0.514572079 0.490849031 0.670600783 0.642611516 PE_(38:5) 0 0.120790615 0.09944068 0.11940586 0.055139892 0.069485408 0.069034835 0.1464627 0.141375402 0.130016357 0.140481364 0.146504362 0.132218098 0.033140331 0.031302044 0.026472523 0.118868888 0.088279992 0.099306427 0.077759946 0.080374011 0.083713038 0.109117685 0.121000341 0.101139732 0.093616604 0.083082828 0.101610902 0.038249148 0.063712982 0.059589718 PE_(38:6) 13043 0.061843457 0.052660812 0.060190642 0.038969007 0.04492237 0.053974707 0.066030571 0.069078255 0.069688517 0.070568477 0.072004059 0.068522475 0.01604715 0.017494728 0.014199032 0.017140986 0.01249462 0.015932785 0.01887232 0.020865083 0.019252194 0.020398069 0.021169293 0.020589508 0.015236297 0.012668607 0.015338109 0.013674158 0.022581015 0.020252755 MS_METABOLITE_DATA_END #METABOLITES METABOLITES_START metabolite_name m/z [M+H]+ Average Rt(min) PE_(34:1) 718.54004 4.781 PE_(34:2) 716.52209 4.303 PE_(35:1) 732.55115 5.03 PE_(36:1) 746.56958 5.243 PE_(36:2) 744.552 4.827 PE_(36:3) 742.5318 4.526 PE_(36:4) 740.52222 4.294 PE_(36:5) 738.50513 3.708 PE_(38:3) 770.56818 5.035 PE_(38:4) 768.55334 4.836 PE_(38:5) 766.53131 4.346 PE_(38:6) 764.52179 4.145 METABOLITES_END #END