PXD033719

PXD033719 is an original dataset announced via ProteomeXchange.

Title	Effects of age and sex on protein abundance in the mouse heart
Description	We compared protein abundance in the left and right ventricles of male and female C57BL6/J mice at 4 months and 20 months old. Frozen tissues were weighed, thawed, and cut into small pieces (2-3 mm3) on weigh boats on ice. Tissue pieces were transferred to 2.0 mL eppendorf tubes. Cold RIPA (Thermo #89901) was added to tissues at a ratio of 20:1 (20 µL RIPA per 1 mg of tissue) and samples were homogenized as above then sonicated with a handheld sonicator (Fisher #FB120110) at 40% amplitude for 15 cycles (each cycle is 1 s of sonication, followed by 5 s pause; three rounds. The samples were centrifuged at 14,000 × g for 15 min at 4°C, and the supernatants were collected. Soluble protein concentration was measured with the BCA protein assay kit following manufacturer instruction; 100 µg of proteins were digested using a filter-assisted protocol as described (Manza et al., 2005) with a 3-hr LysC digestion and 12-hr trypsin digestion. Peptide digest were tagged with tandem mass tags (TMT; 10-plex reagent, Thermo #90110) following manufacturer’s instructions. Tagged digests within the same block are then mixed and fractionated with the high-pH reversed phase peptide fractionation kit (Thermo #84868). Fractionated peptides are dried and resuspended in 0.1% formic acid (LC-MS grade, thermo #28905) at a final concentration of 0.5 µg/µL. Each peptide fraction (~1.5 µg) was further separated with online low-pH reversed-phase LC (PepMap C18 column, 3-μm particle, 100-Å pore; 75 μm x 150 mm; Thermo Fisher Scientific) via the EASYnLC 1200 system coupled to the Easy-Spray ion source (Thermo Fisher Scientific) at 300 nL/min with a 120 min gradient: 0 -105 min: 0 to 40% B; 105 - 110 min: 40 to 70% B; 110 - 115 min: 70 to 100% B; 115 - 120 min: 100% B (solvent A: 0.1% v/v formic acid; solvent B: 80% v/v acetonitrile; column temperature: 50 °C). Mass spectra were acquired on a Thermo Scientific Q-Exactive HF Orbitrap mass spectrometer with the following settings: polarity: positive; data dependent acquisition (DDA): top 15 ions, MS resolution: 60,000, mass range: 300-1650 m/z; precursor dynamic exclusion: 30s, maximum ion injection time: 20 ms; MS automatic gain control (AGC) target: 3e6. isolation window: 1.4 m/z; stepped normalized collision energy (NCE): 28, 30, 32. MS2 resolution; 60,000; MS2 maximum ion injection time: 100 ms; MS2 AGC target: 2e5.
HostingRepository	jPOST
AnnounceDate	2022-11-08
AnnouncementXML	Submission_2022-11-07_14:40:09.125.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Edward Lau
SpeciesList	scientific name: Mus musculus (Mouse); NCBI TaxID: 10090;
ModificationList	TMT; L-methionine sulfoxide; TMT; S-carboxamidomethyl-L-cysteine
Instrument	Q Exactive

Revision	Datetime	Status	ChangeLog Entry
0	2022-05-05 09:50:14	ID requested
⏵ 1	2022-11-07 14:40:10	announced

Han Y, Wennersten SA, Wright JM, Ludwig RW, Lau E, Lam MPY, Proteogenomics reveals sex-biased aging genes and coordinated splicing in cardiac aging. Am J Physiol Heart Circ Physiol, 323(3):H538-H558(2022) [pubmed]

submitter keyword: sex difference, aging, heart, TMT

Maggie Lam
lab head
Edward Lau
contact affiliation	University of Colorado
dataset submitter

jPOST dataset URI

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