Decorin is one of the simplest cellular or pericellular matrix proteoglycan comprised of a core protein containing a single O-linked dermatan/chondroitin sulfate glycosaminoglycan chain. The GAG is a 22k molecular weight chain biosynthesized in the endoplasmic reticulum and Golgi. Xylose is first added to the Ser residue of the GAGylation site of the core protein, followed by two galactose residues and glucuronate residue resulting in tetrasaccharide linker. The chain is extended through glycosyltransferases and modified by sulfotransferase and C5-epimerase. The recent application of high resolution FT-ICR MS to bikunin suggests that it has a single or small number of defined sequences. On this basis, a similar approach to decorin GAG was undertaken, Since decorin is much large and more structurally complex DS/CS GAG chain. Size exclusion and strong anion exchange chromatography were applied to get a low molecular weight and medium charge intact chains without digested by enzyme before preparation-gel separation. The fractions were collected and desalted and then applied to FT-orbitrap MS and FT-ICR MS analysis. Combined the disaccharides analysis results and compared to the theoretical database, we can determined the chain lengths and sulfation patters. The sulfation positions are determined by tandem FT MS. Due to the limitation of MS, we can only detect the chain molecular weight less than 10k, from dp20-dp40 with 7-20 sulfations. The data obtained in this study give us a intact sequence of a relative small chains from the non-reducing end.

Sequence determination of decorin glycosaminoglycan chains

254th National Meeting (2017)

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