#METABOLOMICS WORKBENCH chrisbarlow_20200403_021138 DATATRACK_ID:1979 STUDY_ID:ST001347 ANALYSIS_ID:AN002241 PROJECT_ID:PR000918 VERSION 1 CREATED_ON April 5, 2020, 5:40 pm #PROJECT PR:PROJECT_TITLE Restriction of essential amino acids dictates the systemic metabolic response to PR:PROJECT_TITLE dietary protein dilution PR:PROJECT_SUMMARY The data provided here are in support of the publication “Restriction of PR:PROJECT_SUMMARY essential amino acids dictates the systemic metabolic response to dietary PR:PROJECT_SUMMARY protein dilution” Yann W. Yap et al. Nature Communications 2020 (accepted for PR:PROJECT_SUMMARY publication). Here we provide untargeted metabolomics LC-MS data from liver and PR:PROJECT_SUMMARY plasma from mice fed a diet in which dietary protein was restricted and PR:PROJECT_SUMMARY corresponding unrestricted controls. Specifically, part 1 and 2 correspond to PR:PROJECT_SUMMARY liver and plasma from the hepatic portal vein respectively from animals on a PR:PROJECT_SUMMARY low-protein diet following a week of diet adaptation and correspond controls PR:PROJECT_SUMMARY with n = 5 for each group. Part 3 of the data presented here corresponds to PR:PROJECT_SUMMARY liver from mice subject to a 3wk treatment with diets either containing 18% PR:PROJECT_SUMMARY digestible energy from amino acids or a diet with restricted amounts of PR:PROJECT_SUMMARY threonine and following prior treatments with adeno-associated viruses to PR:PROJECT_SUMMARY transduce the liver to express yeast threonine biosynthetic enzymes PR:PROJECT_SUMMARY (AAV-yTHR1+THR4) or a negative control (AAV-GFP). n= 6 individual mice per PR:PROJECT_SUMMARY group. PR:INSTITUTE Monash University PR:DEPARTMENT Department of Biochemistry and Molecular Biology PR:LABORATORY Nutrient Metabolism and Signalling Laboratory PR:LAST_NAME Rose PR:FIRST_NAME Adam PR:ADDRESS Monash University, Clayton Campus, 23 Innovation Walk, Clayton, Victoria, 3800, PR:ADDRESS Australia PR:EMAIL adam.rose@monash.edu PR:PHONE +61 3 99029340 #STUDY ST:STUDY_TITLE Metabolomic changes in mouse plasma on a protein restricted diet ST:STUDY_SUMMARY The data provided here are in support of the publication “Restriction of ST:STUDY_SUMMARY essential amino acids dictates the systemic metabolic response to dietary ST:STUDY_SUMMARY protein dilution” Yann W. Yap et al. Nature Communications 2020 (accepted for ST:STUDY_SUMMARY publication). Here we provide untargeted metabolomics LC-MS data from plasma ST:STUDY_SUMMARY from C57Bl/6NCrl mice fed a diet in which dietary protein was restricted and ST:STUDY_SUMMARY corresponding unrestricted controls. Specifically, plasma from animals on a ST:STUDY_SUMMARY low-protein diet following a week of diet adaptation and correspond controls ST:STUDY_SUMMARY with n = 5 for each group. ST:INSTITUTE Monash University ST:DEPARTMENT Department of Biochemistry and Molecular Biology ST:LABORATORY Nutrient Metabolism and Signalling Laboratory ST:LAST_NAME Rose ST:FIRST_NAME Adam ST:ADDRESS Monash University, Clayton Campus, 23 Innovation Walk, Clayton, Victoria, 3800, ST:ADDRESS Australia ST:EMAIL adam.rose@monash.edu ST:PHONE +61 3 99029340 ST:NUM_GROUPS 2 ST:TOTAL_SUBJECTS 10 ST:NUM_MALES 10 ST:STUDY_COMMENTS Study 2 for datatrack 1972 #SUBJECT SU:SUBJECT_TYPE Mammal SU:SUBJECT_SPECIES Mus musculus SU:TAXONOMY_ID 10090 SU:GENOTYPE_STRAIN C57Bl/6NCrl SU:AGE_OR_AGE_RANGE 7 to 10 weeks SU:GENDER Male SU:ANIMAL_ANIMAL_SUPPLIER Charles River Laboratories, DEU SU:ANIMAL_LIGHT_CYCLE 12-12h light-dark cycle #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 - AR121_1-2 Diet:Control (normal protein diet) D12450B RAW_FILE_NAME=C120180705_AR121_1-2_1.raw SUBJECT_SAMPLE_FACTORS - AR122_1-3 Diet:Control (normal protein diet) D12450B RAW_FILE_NAME=C120180705_AR122_1-3_1.raw SUBJECT_SAMPLE_FACTORS - AR123_2-1 Diet:Control (normal protein diet) D12450B RAW_FILE_NAME=C120180705_AR123_2-1_1.raw SUBJECT_SAMPLE_FACTORS - AR124_2-2 Diet:Control (normal protein diet) D12450B RAW_FILE_NAME=C120180705_AR124_2-2_1.raw SUBJECT_SAMPLE_FACTORS - AR125_2-3 Diet:Control (normal protein diet) D12450B RAW_FILE_NAME=C120180705_AR125_2-3_1.raw SUBJECT_SAMPLE_FACTORS - AR126_3-2 Diet:Reduced protein diet D10062201 RAW_FILE_NAME=C120180705_AR126_3-2_1.raw SUBJECT_SAMPLE_FACTORS - AR127_3-3 Diet:Reduced protein diet D10062201 RAW_FILE_NAME=C120180705_AR127_3-3_1.raw SUBJECT_SAMPLE_FACTORS - AR128_4-1 Diet:Reduced protein diet D10062201 RAW_FILE_NAME=C120180705_AR128_4-1_1.raw SUBJECT_SAMPLE_FACTORS - AR129_4-2 Diet:Reduced protein diet D10062201 RAW_FILE_NAME=C120180705_AR129_4-2_1.raw SUBJECT_SAMPLE_FACTORS - AR130_4-3 Diet:Reduced protein diet D10062201 RAW_FILE_NAME=C120180705_AR130_4-3_1.raw #COLLECTION CO:COLLECTION_SUMMARY Mice were fasted overnight and then re-fed for 5 to 6 hours before blood was CO:COLLECTION_SUMMARY collected from the hepatic portal vein. CO:SAMPLE_TYPE Blood (plasma) CO:STORAGE_CONDITIONS -80℃ #TREATMENT TR:TREATMENT_SUMMARY Male mice aged 7 weeks upon arrival, were acclimated to the local housing TR:TREATMENT_SUMMARY facility (12-12h light-dark cycle, 22-24°C) for one week prior to TR:TREATMENT_SUMMARY experimentation and were fed standard rodent chow (3437, Provimi Kliba, DEU). TR:TREATMENT_SUMMARY Following acclimation, mice were placed on either a normal protein diet TR:TREATMENT_SUMMARY (D12450B, Research Diets, New Jersey, USA) or a reduced protein diet (D10062201, TR:TREATMENT_SUMMARY Research Diets, New Jersey, USA) for 3 weeks prior to sample collections and TR:TREATMENT_SUMMARY being humanely euthanised for tissue collection. #SAMPLEPREP SP:SAMPLEPREP_SUMMARY Plasma (25 µL) was extracted by addition of 200 µL of 1:1 acetonitrile:MeOH SP:SAMPLEPREP_SUMMARY v/v with 1 µM CHAPS, CAPS, PIPES and TRIS as internal standards at 0°C. SP:SAMPLEPREP_SUMMARY Samples were then mixed on a vortex mixer for 30 minutes at 4°C after which SP:SAMPLEPREP_SUMMARY they were centrifuged (20,000 rcf, 4℃, 15 min) and the supernatant then SP:SAMPLEPREP_SUMMARY transferred to a mass spectrometry vial for LC-MS analysis. SP:PROCESSING_STORAGE_CONDITIONS Described in summary SP:EXTRACT_STORAGE -80℃ #CHROMATOGRAPHY CH:CHROMATOGRAPHY_SUMMARY Chromatography utilized a ZIC-pHILIC column (column temperature 25 °C) with a CH:CHROMATOGRAPHY_SUMMARY gradient elution of 20 mM ammonium carbonate (A) and acetonitrile (B) (linear CH:CHROMATOGRAPHY_SUMMARY gradient time-%B as follows: 0 min-80%, 15 min-50%, 18 min-5%, 21 min-5%, 24 CH:CHROMATOGRAPHY_SUMMARY min-80%, 32 min-80%) on a Dionex RSLC3000 UHPLC (Thermo). The flow rate was CH:CHROMATOGRAPHY_SUMMARY maintained at 300 μL/min. Samples were kept at 4 °C in the autosampler and 10 CH:CHROMATOGRAPHY_SUMMARY μL injected for analysis. CH:CHROMATOGRAPHY_TYPE HILIC CH:INSTRUMENT_NAME Thermo Dionex Ultimate 3000 RS CH:COLUMN_NAME SeQuant ZIC- pHILIC (150 x 4.6 mm, 5uM) #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 spectrometry was performed at 35 000 resolution (accuracy calibrated to <1 MS:MS_COMMENTS ppm) on a Q-Exactive Orbitrap MS (Thermo) operating in rapid switching positive MS:MS_COMMENTS (4 kV) and negative (−3.5 kV) mode electrospray ionization (capillary MS:MS_COMMENTS temperature 300 °C; sheath gas 50; Aux gas 20; sweep gas 2; probe temp 120 MS:MS_COMMENTS °C). MS:MS_RESULTS_FILE ST001347_AN002241_Results.txt UNITS:Peak Area Has m/z:Neutral masses Has RT:Yes RT units:Minutes #END