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Determination of sulfation pattern in brain glycosaminoglycans by chip-based electrospray ionization ion trap mass spectrometry

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Abstract

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 (MS2–MS3) 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 β-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-Δ-[IdoA-GalNAc]. By optimized CID MS2–MS3, 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-Δ-[GlcA-GalNAc]. The site of oversulfation was determined by MS2–MS3, which provided sequence patterns consistent with a rare GlcA-3-sulfate–GalNAc-6-sulfate structural motif.

Mouse brain GlcA-3-sulfate-GalNAc-6-sulfate structural motif

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Acknowledgments

We are grateful to Prof. Dr. Ecaterina Andronescu, Romanian Minister of Education, Research and Innovation, and Prof. Dr. Lizica Mihut, Rector of “Aurel Vlaicu” University of Arad, Romania, for their invaluable support.

This work was supported by Romanian National Authority for Scientific Research through the grants CE.EX. 111/2006, 98/2006, and PN-II-41001/2007 and German Society for Research (grant SFB 492/A9 and DFG SE1431/1-1).

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Authors and Affiliations

  1. Mass Spectrometry Laboratory, National Institute for Research and Development in Electrochemistry and Condensed Matter, Plautius Andronescu Str. 1, 300224, Timisoara, Romania

    Corina Flangea, Catalin Schiopu, Alina Serb & Alina D. Zamfir

  2. “Victor Babes” University of Medicine and Pharmacy, Eftimie Murgu Square, 2, 300054, Timisoara, Romania

    Corina Flangea, Eugen Sisu & Alina Serb

  3. Chemistry Institute of Romanian Academy, Mihai Viteazul Blvd 24, 300223, Timisoara, Romania

    Eugen Sisu

  4. Laboratory of Analytical Chemistry and Biopolymer Structure Analysis, University of Konstanz, Universitätsstrasse 10, 78464, Konstanz, Germany

    Michael Przybylski

  5. Institute for Physiological Chemistry and Pathobiochemistry, University of Münster, Waldayer Str. 15, 48149, Münster, Germany

    Daniela G. Seidler

  6. Department of Chemical and Biological Sciences, “Aurel Vlaicu” University of Arad, Revolutiei Blvd 77, 310130, Arad, Romania

    Alina D. Zamfir

Authors
  1. Corina Flangea
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  2. Catalin Schiopu
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  3. Eugen Sisu
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  4. Alina Serb
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  5. Michael Przybylski
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  6. Daniela G. Seidler
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  7. Alina D. Zamfir
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Correspondence to Alina D. Zamfir.

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Flangea, C., Schiopu, C., Sisu, E. et al. Determination of sulfation pattern in brain glycosaminoglycans by chip-based electrospray ionization ion trap mass spectrometry. Anal Bioanal Chem 395, 2489–2498 (2009). https://doi.org/10.1007/s00216-009-3167-0

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  • DOI: https://doi.org/10.1007/s00216-009-3167-0

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