Dataset Information

A workflow towards the reproducible identification and quantitation of protein carbonylation sites in human plasma

ABSTRACT: Protein carbonylation has been consistently used as a marker of excessive oxidative stress and studied in the context of multiple human oxidative stress related diseases. The variety of carbonyl post-translational modifications (PTMs) and their low abundance on easily accessible diagnostic tissues (e.g., plasma) challenges their reproducible identification and quantitation. However, the use of carbonyl-specific biotinylated derivatization tags (e.g., aldehyde reactive probe, ARP) allows targeting carbonyl PTMs and enriching proteins and/or peptides carrying these modifications that facilitate their characterization. In this study, an oxidized human serum albumin protein model (OxHSA) and plasma proteins from a healthy individual were derivatized with ARP, digested with trypsin, and ARP-derivatized peptides (ARP-peptides) were enriched using biotin-avidin affinity chromatography prior to RPC-TWIMS-MS/MS analysis. The present workflow highlights previously overlooked analytical challenges, shows the use of ARP specific fragment ions to improve identification reliability of ARP-peptide identifications, and displays an extensive validation of the reproducible recovery and relative quantitation of ARP-peptides. HSA was identified as the most heavily modified protein by a variety of direct amino acid oxidation products and adducts from reactive carbonyl species (RCS). Most RCS modification sites were identified in six HSA modification hotspots (Lys10, Lys190, Lys199, Lys281, Lys432, and Lys525).

INSTRUMENT(S): Synapt MS

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Blood Plasma

SUBMITTER: Juan Camilo Rojas Echeverri

LAB HEAD: Ralf Hoffmann

PROVIDER: PXD023738 | Pride | 2021-03-02

REPOSITORIES: Pride

ACCESS DATA

Dataset's files

Source:

			Action	DRS
	190404_Homo-sapiens_Swiss-Prot-uniprot-9606.fasta	Fasta
	ARP-OxHSA_DilSeries.zip	Other
	ARP-OxHSA_DilSeries_PTM_fix_cbmm_5perc_PepLevel.pep.xml	Pepxml
	ARP-OxHSA_DilSeries_PTM_search_fixed_cbmm_5perc_PepLevel_psm.csv	Csv
	ARP-Pl-OxHSA_DB_fixed_cbmm.mzid.gz	Mzid

Items per page:

1 - 5 of 133

Similar Datasets

Project description:MarekM-bM-^@M-^Ys disease virus (MDV) is an oncovirus causing tumor disease known as MarekM-bM-^@M-^Ys disease (MD) in chicken. Breeding of chickens genetically resistant to MD is considered a vital augment to better control MD. To find the mechanism underlying the genetic resistance to MD, a genomic structural variation, copy number variation (CNV), was examined in inbred MD-resistant and -susceptible chicken lines by using the comparative genomic hybridization (CGH) technique. A total of 45 copy number variation regions (CNVRs) were found spanning across 3,297,038 bp in length of the chicken genome in 4 lines of chickens. Ten CNVRs were selectively confirmed with quantitative real-time PCR. The comparison between the resistant and susceptible chicken lines revealed 28 differentially presented CNVRs, which are functionally involved in immune response, cell proliferation in midbrain, G-protein coupled receptor signaling pathway, and protein-glutamine gamma-glutamyltransferase activity. Two CNVRs that are related with MD-resistance and M-bM-^@M-^Ssusceptibility were also found transmitted to descendent recombinant congenic lines that differ in susceptibility to MD. A positive correlation was identified between the CNVRs and gene expression, indicating the importance of gene expression dosage in disease resistance. We also found the overlapping between the CNVR region and the MarekM-bM-^@M-^Ys disease trait related quantitative trait loci (QTLs). In conclusion, our data provided additional information elucidating one of possible mechanisms underlying of genetic resistance to MD. The findings may eventually lead to better strategies for genetic improvement of resistance to MD in poultry. L63: highly resistant to MD; L72: highly susceptible to MD; RCS-L: moderate resistant to MD; RCS-M: moderate susceptible to MD Red blood cell DNAs were isolated from L63, L72, RCS-L and RCS-M and then labeled with Cy3. DNA from Red Junglefowl was labeled with Cy5 as reference and mixed with Cy3 labeled DNA, then hybridized to the 385K chicken tiling array. The hybridization, normalization and segmentation analysis were performed by NimbleGen Systems Inc. (Madison, USA). A detailed description of their technical specifics can be found on http://www.nimblegen.com/products/lit/lit.html. Briefly, the segments with five or more probes having a mean log2 ratio greater than M-BM-10.5 (0.5_5) were chosen as candidate CNVs.

Project description:The presentation of virus-derived peptides by HLA class I molecules on the surface of an infected cell and the recognition of these HLA-peptide complexes by, and subsequent activation of, CD8+ cytotoxic T cells provides an important mechanism for immune protection against viruses. Recent advances in proteogenomics have allowed researchers to discover a growing number of unique HLA-restricted viral peptides, resulting in a rapidly expanding repertoire of targets for immunotherapeutics (i.e. bispecific antibodies, engineered T-cell receptors (TCRs), chimeric antigen receptor T-cells (CAR-Ts)) to infected tissues. However, genomic variability between viral strains, such as Hepatitis-B virus (HBV), in combination with differences in patient HLA alleles, make it difficult to develop therapeutics against these targets. To address this challenge, we developed a novel proteogenomics approach for generating patient-specific databases that enable the identification of viral peptides based on the viral transcriptomes sequenced from individual patient liver samples. We also utilized DNA sequencing of patient samples to identify HLA genotypes and assist in target selection. Liver samples from 48 HBV infected patients, primarily from Asia, were examined to reconstruct patient-specific HBV genomes, identify regions within the human chromosomes targeted by HBV integrations and obtain a comprehensive view of HBV peptide epitopes using our HLA class-I (HLA-I) immunopeptidomics discovery platform. Two previously reported HLA associated HBV-derived peptides, HLA-A02 binder FLLTRILTI (S194-202) from the large surface antigen and HLA-A11 binder STLPETTVVRR (C141-151) from the capsid protein were validated by our discovery platform, but both were detected at a very low frequencies. In addition, we identified and validated, using heavy peptide analogues, novel strain-specific HBV-HLA associated peptides, such as GSLPQEHIVQK (P606-616) and variants. Overall, our novel approach can guide the development of bispecific antibody, TCR-T, or CAR-T based therapeutics for the treatment of HBV-related HCC and inform vaccine development.

Dataset Information

A workflow towards the reproducible identification and quantitation of protein carbonylation sites in human plasma

Dataset's files

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets