Determination of sulfation pattern in brain glycosaminoglycans by chip-based electrospray ionization ion trap mass spectrometry.
Chondroitin sulfate (CS) and dermatan sulfate (DS) glycosaminoglycans display variability of sulfation in their constituent disaccharide repeats during chain elongation. Since a large proportion of the extracellular matrix of the central nervous system (CNS) is composed of proteoglycans, CS/DS disaccharide degree and profile of sulfation play important roles in the functional diversity of neurons, brain development, and some of its pathological states. To investigate the sulfation pattern of CS/DS structures expressed in CNS, we introduced here a novel method based on an advanced system encompassing fully automated chip nanoelectrospray ionization (nanoESI) in the negative ion mode and high capacity ion trap multistage mass spectrometry (MS(2)-MS(3)) by collision-induced dissociation (CID). This method, introduced here for the first time in glycomics of brain glycosaminoglycans, was particularly applied to structural investigation of disaccharides obtained by beta-elimination and digestion with chondroitin B and AC I lyase of hybrid CS/DS chains from wild-type mouse brain. Screening in the chip-MS mode of DS disaccharide fraction resulting after depolymerization with chondroitin B lyase revealed molecular ions assigned to monosulfated disaccharide species having a composition of 4,5-Delta-[IdoA-GalNAc]. By optimized CID MS(2)-MS(3), fragment ions supporting the localization of sulfate ester group at C4 within GalNAc were produced. Chip ESI MS profiling of CS disaccharide fraction obtained by depolymerization of the same CS/DS chain using chondroitin AC I lyase indicated the occurrence of mono- and bisulfated 4,5-Delta-[GlcA-GalNAc]. The site of oversulfation was determined by MS(2)-MS(3), which provided sequence patterns consistent with a rare GlcA-3-sulfate-GalNAc-6-sulfate structural motif. Figure Mouse brain GlcA-3-sulfate-GalNAc-6-sulfate structural motif.
Flangea
Corina
C
Mass Spectrometry Laboratory, National Institute for Research and Development in Electrochemistry and Condensed Matter, Plautius Andronescu Str. 1, 300224, Timisoara, Romania.
Schiopu
Catalin
C
Sisu
Eugen
E
Serb
Alina
A
Przybylski
Michael
M
Seidler
Daniela G
DG
Zamfir
Alina D
AD
D000818:Animals; D001921:Brain / Q000378:metabolism*; D002809:Chondroitin Sulfates / Q000032:analysis*; D003871:Dermatan Sulfate / Q000032:analysis*; D004187:Disaccharides / Q000032:analysis; D006025:Glycosaminoglycans / Q000737:chemistry* / Q000378:metabolism*; D051379:Mice; D008810:Mice, Inbred C57BL; D036103:Nanotechnology; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*; D013431:Sulfates / Q000032:analysis*
D004187:Disaccharides; D006025:Glycosaminoglycans; D013431:Sulfates; D003871:Dermatan Sulfate; D002809:Chondroitin Sulfates