Dataset Information

LC-MS/MS Isomeric Profiling of N-Glycans derived from Low-Abundant Serum Glycoproteins in Mild Cognitive Im-pairment Patients

ABSTRACT: Mild cognitive impairment (MCI) is an early stage of memory loss that affects cognitive abilities, such as language or virtual/spatial comprehension. This cognitive decline is mostly ob-served with the aging of individuals. Recently, MCI has been considered as a prodromal phase of Alzheimer’s disease (AD), with a 10-15% conversion rate. However, the existing diagnostic methods fail to provide precise and well-timed diagnoses, and the pathophysiology of MCI is not fully understood. Alterations of serum N-glycan expression could represent essential con-tributors to the overall pathophysiology of neurodegenerative diseases and be used as a poten-tial marker to assess MCI diagnosis using non-invasive procedures. Herein, we undertook an LC-MS/MS glycomics approach to determine and characterize potential N-glycan markers in de-pleted blood serum samples from MCI patients. For the first time, we profiled the isomeric gly-come of the low abundant serum glycoproteins extracted from serum samples of control and MCI patients using an LC-MS/MS analytical strategy. Additionally, the MRM validation of the identified data showed five isomeric N-glycans with the ability to discriminate between healthy and MCI patients: the sialylated N-glycans HexNAc5,Hex6,Neu5Ac3 and HexNAc6,Hex7,Neu5Ac4 with single AUCs of 0.92 and 0.87 respectively, and combined AUC of 0.96; and the sialylat-ed-fucosylated N-glycans HexNAc4,Hex5,Fuc,Neu5Ac, HexNAc5,Hex6,Fuc,Neu5Ac2 and HexNAc6,Hex7,Fuc,Neu5Ac3 with single AUCs of 0.94, 0.67, and 0.88 respectively, and combined AUC of 0.98. According to ingenuity pathway analysis (IPA) and in line with recent publications the identified N-glycans may play an important role in neuroinflammation. Process that plays a fundamental role in the progression of neurodegenerative diseases.

INSTRUMENT(S): LTQ Orbitrap Velos, TSQ Vantage

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Blood Cell

DISEASE(S): Mild Cognitive Impairment

SUBMITTER: Cristian Gutierrez Reyes

LAB HEAD: Yehia Mechref

PROVIDER: PXD036704 | Pride | 2023-03-11

REPOSITORIES: Pride

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Publications

LC-MS/MS Isomeric Profiling of <i>N</i>-Glycans Derived from Low-Abundant Serum Glycoproteins in Mild Cognitive Impairment Patients.

Reyes Cristian D Gutierrez CDG Hakim Md Abdul MA Atashi Mojgan M Goli Mona M Gautam Sakshi S Wang Junyao J Bennett Andrew I AI Zhu Jianhui J Lubman David M DM Mechref Yehia Y

Biomolecules 20221108 11

Mild cognitive impairment (MCI) is an early stage of memory loss that affects cognitive abilities, such as language or virtual/spatial comprehension. This cognitive decline is mostly observed with the aging of individuals. Recently, MCI has been considered as a prodromal phase of Alzheimer's disease (AD), with a 10-15% conversion rate. However, the existing diagnostic methods fail to provide precise and well-timed diagnoses, and the pathophysiology of MCI is not fully understood. Alterations of ...[more]

PMID: 36359007

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Project description:Alterations in glycosylation are seen in many types of cancer, including colorectal cancer (CRC). CRC is known to display glycosylation alterations. Glycans, the sugar moieties of glycoconjugates, are involved in many important functions relevant to cancer, such as cell signaling and adhesion, and may can be of value as biomarkers. In this study, we have used mass spectrometry to analyze the N-glycan profiles of 35 CRC tissue samples from patients with tumors in the right or left colon and 10 healthy tissue samples from non-CRC patients who underwent operations for other reasons. The tumor samples were divided into groups depending on tumor location (right or left colon) and stage (II or III), while the healthy samples were divided into right or left side of the colon. The levels of neutral and acidic N-glycan compositions and glycan classes were analyzed in a total of ten different groups. Surprisingly, there were no significant differences in glycan levels when all right- and left-sided CRC samples were compared, and few differences (such as in the abundance of the neutral N-glycan H3N5) were seen when the samples were divided according to both location and stage. Multiple significant differences were found in the levels of glycans and glycan classes when stage II and III samples were compared, and these glycans could be of value as candidates for new markers of cancer progression. In order to validate our findings, we analyzed healthy tissue samples from the right and left colon and found no significant differences in the levels of any of the glycans analyzed, confirming that our findings when comparing CRC samples from the right and left colon are not due to normal variations in the levels of glycans between the healthy right and left colon. Additionally, the levels of the acidic glycans H4N3F1P1, H5N4F1P1, and S1H5N4F1 were found to change in a cancer-specific but colon location-nonspecific manner, indicating that CRC affects glycan levels in similar ways, regardless of tumor location.

Project description:Recent advances in software-driven glycopeptide identification in LC-MS/MS-based N-glycoproteomics have facilitated biochemical studies reporting thousands of intact N-glycopeptides, i.e. N-glycan-conjugated peptides, but the automated identification process remains to be scrutinized. Herein, we explore the efficiency of site-specific glycoprofiling using the PTM-centric search-engine Byonic relative to manual expert annotation. To allow an appropriately deep comparison, the study utilised typical glycoproteomics acquisition and data analysis strategies, but of a single glycoprotein, the uncharacterised N-glycosylated (Asn160, Asn268 and Asn302) human basigin. Detailed site-specific reference glycoprofiles of purified basigin were manually established using ion trap CID-MS/MS and high-resolution Q-Exactive Orbitrap HCD-MS/MS acquisition of tryptic N-glycopeptides and released N-glycans. The basigin N-glycosylation sites, which showed extensive micro- and macro-heterogeneity, were then glycoprofiled using Byonic with or without a background of complex peptides using Q-Exactive Orbitrap HCD-MS/MS data. The glycoprofiling efficiencies were assessed against the site-specific reference glycoprofiles and target and decoy proteome databases. The search criteria and confidence thresholds (Byonic scores) recommended by the vendor provided very high glycoprofiling accuracy and coverage (both >80%) and low peptide FDRs (<1%). The data complexity, search parameters including search space (proteome/glycome size), mass tolerance and peptide modifications, and confidence thresholds affected the glycoprofiling efficiency and analysis time. Automated identification of peptide modifications (methionine oxidation/carbamidomethylation) that coincide with monosaccharide mass differences (Fuc/Hex/HexNAc) and accurately distinguishing isobaric (Hex1NeuAc1-R/Fuc1NeuGc1-R) or near-isobaric (NeuAc1-R/Fuc2-R) monosaccharide sub-compositions remain challenging, arguing particular attention to such “difficult-to-identify” N-glycopeptides. The presented analysis provides valuable insights into automated glycopeptide identification; knowledge that facilitates further developments in FDR-based glycoproteomics.

Dataset Information

LC-MS/MS Isomeric Profiling of N-Glycans derived from Low-Abundant Serum Glycoproteins in Mild Cognitive Im-pairment Patients

Publications

LC-MS/MS Isomeric Profiling of <i>N</i>-Glycans Derived from Low-Abundant Serum Glycoproteins in Mild Cognitive Impairment Patients.

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