Extension of the in-gel release method for structural analysis of neutral and sialylated N-linked glycans to the analysis of sulfated glycans: application to the glycans from bovine thyroid-stimulating hormone.
This paper reports an extension of the in-gel technique for releasing N-linked glycans from glycoproteins for analysis by matrix-assisted laser desorption/ionization (MALDI) mass spectrometry reported by B. Küster, S. F. Wheeler, A. P. Hunter, R. A. Dwek, and D. J. Harvey (1997, Anal. Biochem. 250, 82-101) to allow it to be used for sulfated glycans. The method was used to identify N-linked glycans from bovine thyroid-stimulating hormone. Following glycan release, either in gel or in solution, the glycans were fractionated directly with a porous graphatized carbon column. The sulfated glycans were examined by MALDI mass spectrometry in negative ion mode with d-arabinosazone as the matrix and both neutral and acidic glycans were examined in positive ion mode from 2,5-dihydroxybenzoic acid. Negative ion post-source decay spectra were also obtained. Twenty-two neutral and fifteen sulfated N-linked glycans were identified and it was shown that negligible loss of sulfate groups occurred even though these groups are often readily lost during MALDI analysis. The glycans were mainly sulfated hybrid and biantennary complex structures. Negative ion post-source decay and positive ion collision-induced fragmentation spectra were obtained.
Wheeler
S F
SF
Department of Biochemistry, Oxford Glycobiology Institute, South Parks Road, Oxford, OX1 3QU, United Kingdom.
Harvey
D J
DJ
D000581:Amidohydrolases / Q000378:metabolism; D000818:Animals; D002417:Cattle; D004591:Electrophoresis, Polyacrylamide Gel; D043524:Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase; D011134:Polysaccharides / Q000032:analysis* / Q000737:chemistry* / Q000378:metabolism; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013972:Thyrotropin / Q000737:chemistry*
D011134:Polysaccharides; D013972:Thyrotropin; D000581:Amidohydrolases; D043524:Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase