Glycation pattern of peptides condensed with maltose, lactose and glucose determined by ultraviolet matrix-assisted laser desorption/ionization tandem mass spectrometry

Abstract

Protein glycation is the non-enzymatic condensation of sugars with proteins. Although commonly occurring in both the therapeutic and food/beverage industries, protein glycation has not been the focus of many proteomic investigations. This study aims to establish a reliable mass spectrometric method for screening large tandem mass spectrometric (MSMS) datasets for protein glycation with glucose, lactose and maltose. Control experiments using a standard peptide containing a single glycation site led to the discovery of characteristic neutral loss fragmentation patterns in MSMS analysis for glucose, lactose and maltose condensed with peptides. Valid in both tandem time-of-flight (TOFTOF) and quadrupole ion trap time-of-flight matrix-assisted laser desorption/ionization (QIT TOF MALDI) mass spectrometers, these neutral loss signatures were then applied to elucidation of modified peptides from a complex human serum albumin (HSA) digest glycated with each of the proposed sugars. Screening of these large datasets was made possible by specifically designed software solutions that enable the input of detailed user-defined post-translational modifications that are not included in the universally available databases such as Unimod.

Rapid Fc glycosylation analysis of Fc fusions with IdeS and liquid chromatography mass spectrometry

“…Maltose glycation on proteins through lysine has been observed. 16 One maltose added 324 Da to the glycated protein, which was the exact mass addition we observed here. It was reasonable for us to believe that the observed 324 Da modification was a maltose glycation that resulted from the storage of reconstituted abatacept powder formulated with maltose.…”

Non‐enzymatic glycation and glycoxidation protein products in foods and diseases: An interconnected, complex scenario fully open to innovative proteomic studies

“…However, CID experiments on glucosylated peptides revealed that sequence‐informative b and y ions resulting from the peptide backbone cleavage were reduced within MS/MS spectra, hampering sequence‐derived peptide identification. In the case of glucosylated peptides, the most abundant ions corresponded to the NL of H 2 O (−18 Da), 2 H 2 O (−36 Da), 3 H 2 O (−54 Da), 4 H 2 O (−72 Da), 3 H 2 O + HCHO (−84 Da), and of the Amadori adduct, C 6 H 10 O 5 (−162 Da) (Zhang et al, ; Lapolla et al, ; Brancia et al, ; Frolov, Hoffmann, & Hoffmann, ; Gadgil et al, ; Montgomery, Tanaka, & Belgacem, ; Guedes et al, ; Chougale et al, ); consecutive losses of H 2 O, 3 H 2 O, and 3 H 2 O + HCHO yield oxonium, pyrylium, and furylium ions, respectively. When MALDI‐TOF instrument were used, NL of C 4 H 8 O 4 (−120 Da) was also detected, which originates by the cleavage of the glycan moiety (Brancia et al, ; Montgomery, Tanaka, & Belgacem, ).…”

Analysis of protein glycation using fluorescent phenylboronate gel electrophoresis

“…BAC has been successfully used as a sample enrichment step prior to high-performance liquid chromatography (HPLC) and mass spectrometry (MS) analyses14. Even though MS15 is an ideal technique to determine the identity of protein band in a gel, the limited availability of database information specific for glycation adducts hampers the identification of the modifications. In addition, MS only detects the most abundant proteins and if only a small percentage of a given protein species in a sample has suffered glycation damage, these adducts are likely to go undetected.…”