Mass spectrometry dataset on apo-SOD1 modifications induced by lipid aldehydes

Metal-deficient Cu,Zn-superoxide dismutase (apo-SOD1) is associated with the formation of SOD1 aggregates that accumulate in ALS disease. The data supplied in this article support the accompanying publication showing SOD1 modification and aggregation induced by lipid aldehydes [1]. Here, we present the LC-MS/MS dataset on apo-SOD1 modification induced by seven different lipid aldehydes: 4-hydroxy-2-hexenal (HHE), 4-hydroxy-2-nonenal (HNE), 2-hexen-1-al (HEX), 2,4-nonadienal (NON), 2,4-decadienal (DEC) or secosterol aldehydes (SECO-A or SECO-B). Modified protein samples were digested with trypsin and sequenced by a LC coupled to a Q-TOF instrument. Protein sequencing and peptide modification analysis was performed by Mascot 2.6 (Matrix Science) and further validated by manual inspection. Mass spectrometry data (RAW files) obtained in this study have been deposited to MassIVE and the observed peptide-aldehyde adducts can be used in further studies exploring SOD1 modifications in vivo.


a b s t r a c t
Metal-deficient Cu,Zn-superoxide dismutase (apo-SOD1) is associated with the formation of SOD1 aggregates that accumulate in ALS disease. The data supplied in this article support the accompanying publication showing SOD1 modification and aggregation induced by lipid aldehydes [1] . Here, we present the LC-MS/MS dataset on apo-SOD1 modification induced by seven different lipid aldehydes: 4-hydroxy-2-hexenal (HHE), 4-hydroxy-2-nonenal (HNE), 2-hexen-1-al (HEX), 2,4-nonadienal (NON), 2,4-decadienal (DEC) or secosterol aldehydes (SECO-A or SECO-B). Modified protein samples were digested with trypsin and sequenced by a LC coupled to a Q-TOF instrument. Protein sequencing and peptide modification analysis was performed by Mascot 2.6 (Matrix Science) and further validated by manual inspection. Mass spectrometry data (RAW files) obtained in this study have been deposited to MassIVE and the observed peptidealdehyde adducts can be used in further studies exploring SOD1 modifications in vivo .
© 2020 The Author(s

Value of the data
• The data show the characterization of apo-SOD1 lipoxidation sites induced by seven biologically relevant lipid aldehydes. • These data can be useful for researchers studying protein lipoxidation.
• These data can be useful for studies investigating protein post-translational modifications induced by lipid peroxidation products.

Protein digestion
After incubation, samples were first reduced with sodium borohydride (NaBH 4 , 5 mM), for 1 h at room temperature and then, submitted to disulfide reduction with dithiothreitol (DTT, 5 mM), for 30 min at 60 °C and Cys alkylation with iodoacetamide (15 mM), for 30 min at room temperature. Protein digestion was done with proteomic-grade trypsin (Promega, Madison, WI,

LC-MS/MS analysis
Peptide mixture was analyzed by a LC-MS/MS system consisted of a nanoAcquity UPLC system (Waters Corp., Milford, MA, USA), coupled to a quadrupole-time-of-flight (Q-TOF) mass spectrometer (TripleTOF6600 Sciex, United States), as described previously [4] . First, samples were desalted on the trapping column (Waters, nanoAcquity Trap column, 180 μm × 20 mm; 5 μm) using 1% solvent B at a flow rate of 10 μL/min for 2 min under isocratic conditions. Peptides were then separated on a C18 analytical column (Waters nanoAcquity UPLC, 75 μm × 150 mm; 3.5 μm) using a gradient of 0.1% aqueous formic acid (mobile phase A) and 0.1% formic acid in acetonitrile (mobile phase B). Chromatographic separation was done at a flow rate of 400 nL min −1 for a total run time of 97 min according to a gradient shown below. Column temperature was kept at 35 °C. Sample injection volume was 2 μL. Peptides were infused into the TripleTOF6600 instrument through a nano-ESI source (Sciex, Framingham, MA). The nano-ESI source was equipped with a nano-ESI emitter tip (New Objective). The mass spectrometer parameters were:

Ion Source Parameters Settings
Ion spray voltage floating (ISVF) 2400 V Curtain Gas (CUR) 20 Interface heater (IHT) 120 Ion source gas 1 (GS1) 3 Ion source gas 2 (GS2) 0 Declustering potential (DP) 80 V Tandem mass spectra were acquired by a data-dependent mode. TOFMS survey scan was set to the m/z range of 30 0-20 0 0 and the accumulation time to 100 ms. Top 25 MS/MS spectra were acquired in the mass range of m/z 10 0-20 0 0 with an accumulation time of 25 ms. The overall cycle time was 775 ms. Precursor ion selection criteria included charge state between + 2 and + 5 and ion intensity greater than 150 counts. Former fragmented precursor ions were excluded from reanalysis for 20 s. Fragmentation was performed using rolling collision energy with a collision energy spread of 5. For LC-MS/MS quality control we used 1 pmol/μl stock solution of beta-galactosidase, which was prepared according to manufacturer's instruction (LC/MS peptide calibration kit P/N 4,465,867), pre-digested BSA or HeLa protein digest standard (Pierce, Thermo Scientific). Data acquisition was performed with Analyst TF 1.7 (Sciex). Mass spectrometry raw data have been deposited to the Mass Spectrometry Interactive Virtual Environment (MassIVE), with access via https://massive.ucsd.edu/ProteoSAFe/dataset.jsp?accession=MSV0 0 0 085309 .

Data analysis
Protein sequencing and modification analysis was performed with Mascot R software 2.6.1 version (Matrix Science Ltd., London, United Kingdom), using the following parameters: Modified peptides identified by Mascot R were further validated by manual inspection. To identify the y and b fragments of the modified peptides and attribute their masses in MS/MS spectrum, we used the Bio Tool Kit microapp in PeakView R software. First, the protein sequence was digested in silico to create a list of theoretical peptides. Modified SOD1 peptide sequences found in MASCOT R software with their respective charges were selected in "Bio Tool Kit" and modifications corresponding to the mass of each aldehyde were added as variable modification. The software was settled to match the theoretical fragments to the ions in MS/MS spectrum with a match tolerance of 0.050 Da.

Declaration of Competing Interest
The authors declare that they have no competing financial interests or personal relationships that could influence the work reported in this paper.