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Multiplexed glycoproteomic analysis of glycosylation disorders by sequential yolk immunoglobulins immunoseparation and MALDI‐TOF MS
Author(s) -
Sturiale Luisa,
Barone Rita,
Palmigiano Angelo,
Ndosimao Célestin Nsibu,
Briones Paz,
Adamowicz Maciej,
Jaeken Jaak,
Garozzo Domenico
Publication year - 2008
Publication title -
proteomics
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.26
H-Index - 167
eISSN - 1615-9861
pISSN - 1615-9853
DOI - 10.1002/pmic.200700496
Subject(s) - glycosylation , antibody , matrix assisted laser desorption/ionization , yolk , chemistry , proteomics , n linked glycosylation , biochemistry , biology , glycoprotein , glycan , immunology , food science , organic chemistry , adsorption , desorption , gene
This study applied yolk immunoglobulins immunoaffinity separation and MALDI‐TOF MS for clinical proteomics of congenital disorders of glycosylation (CDG) and secondary glycosylation disorders [galactosemia and hereditary fructose intolerance (HFI)]. Serum transferrin (Tf) and α1‐antitrypsin (AAT) that are markers for CDG, were purified sequentially to obtain high‐quality MALDI mass spectra to differentiate single glycoforms of the native intact glycoproteins. The procedure was found feasible for the investigation of protein macroheterogeneity due to glycosylation site underoccupancy then ensuing the characterization of patients with CDG group I (N‐glycan assembly disorders). Following PNGase F digestion of the purified glycoprotein, the characterization of protein microheterogeneity by N‐glycan MS analysis was performed in a patient with CDG group II (processing disorders). CDG‐Ia patients showed a typical profile of underglycosylation where the fully glycosylated glycoforms are always the most abundant present in plasma with lesser amounts of partially and unglycosylated glycoforms in this order. Galactosemia and HFI are potentially fatal diseases, which benefit from early diagnosis and prompt therapeutic intervention. In symptomatic patients with galactosemia and in those with HFI, MALDI MS of Tf and AAT depicts a hypoglycosylation profile with a significant increase of underglycosylated glycoforms that reverses by dietary treatment, representing a clue for diagnosis and treatment monitoring.