Nov 12, 2023
Proteome analysis
- 1University of California, San Diego
Protocol Citation: Leonardo A Parra-Rivas 2023. Proteome analysis. protocols.io https://dx.doi.org/10.17504/protocols.io.5jyl8pw86g2w/v1
License: This is an open access protocol distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited
Protocol status: Working
We use this protocol and it's working
Created: November 12, 2023
Last Modified: November 12, 2023
Protocol Integer ID: 90828
Abstract
Proteome analysis
Mass spectrometry data was
analyzed according to a published protocol
CITATION
Spectra were searched using Proteome Discoverer (RRID:SCR_014477) (https://www.thermofisher.com/order/catalog/product/IQLAAEGABSFAKJMAUH) software version 2.1 against 2020 mouse UniProtKB/Swiss-Prot
(RRID:SCR_021164)( https://www.expasy.org/resources/uniprotkb-swiss-prot) (17,042 target sequences) along with the human α-synuclein protein sequence.
Searching parameters included full tryptic or Asp-N restriction, precursor mass tolerance
(± 20 ppm), and fragment mass tolerance (± 0.05 Da). Serine, threonine, and tyrosine
phosphorylation (+79.9663 Da), methionine oxidation (+15.99492 Da),
asparagine and glutamine deamidation (+0.98402 Da), and protein N-terminal
acetylation (+42.03670 Da) were variable modifications (up to 3 allowed per
peptide); cysteine was assigned a fixed carbamidomethyl modification (+57.021465
Da).
Percolator was used to filter the peptide spectrum matches to a false
discovery rate of 1%.
Gene ontology was analyzed using The Database for Annotation, Visualization and
Integrated Discovery DAVID (RRID:SCR_001881)(https://david.ncifcrf.gov/tools.jsp-DAVID97.
Biological processes involving synaptic function were selected for grouping analysis. Functional grouping was based on Fisher’s exact test (p<0.05).
Protein-protein interaction networks were then identified using the STRING (RRID:SCR_005223 ) (https://string-db.org/) version 11.5 , and the protein class was determined using PANTHER
(RRID:SCR_004869)( http://www.pantherdb.org/).
Citations
Step 1
Wingo TS, Duong DM, Zhou M, Dammer EB, Wu H, Cutler DJ, Lah JJ, Levey AI, Seyfried NT. Integrating Next-Generation Genomic Sequencing and Mass Spectrometry To Estimate Allele-Specific Protein Abundance in Human Brain.
https://doi.org/10.1021/acs.jproteome.7b00324