title: Method development and analysis of free HS and HS in proteoglycans from pre- and postmenopausal women: evidence for biosynthetic pathway changes in sulfotransferase and sulfatase enzymes.
abstract: Heparan sulfate (HS) is one of the most complex and informative biopolymers found on the cell surface or in the extracellular matrix as either free HS fragments or constituents of HS proteoglycans (HSPGs). Analysis of free HS and HSPG sugar chains in human serum at the disaccharide level has great potential for early disease diagnosis and prognosis; however, the low concentration of HS in human serum, together with the complexity of the serum matrix, limits the information on HS. In this study, we present and validate the development of a new sensitive method for in-depth compositional analysis of free HS and HSPG sugar chains. This protocol involved several steps including weak anion exchange chromatography, ultrafiltration, and solid-phase extraction for enhanced detection prior to LC-MS/MS analysis. Using this protocol, a total of 51 serum samples from 26 premenopausal and 25 postmenopausal women were analyzed. Statistically significant differences in heparin/HS disaccharide profiles were observed. The proportion of N-acetylation and N-sulfation in both free HS and HSPG sugar chains were significantly different between pre- and postmenopausal women, indicating changes in N-deacetylase/N-sulfotransferases (NDSTs), the enzymes involved in the initial step of the biosynthetic pathway. Differences in the proportion of 6-O-sulfation suggest that 6-O-sulfotransferase and/or 6-O-sulfatase enzymes may also be implicated.
authors: Department of Chemistry, University of California, Davis, California 95616, USA.
authors: Wei
authors: W
authors: Wei
authors: Rebecca L
authors: RL
authors: Miller
authors: Julie A
authors: JA
authors: Leary
mesh_terms: D000328:Adult; D000368:Aged; D053898:Biosynthetic Pathways / Q000502:physiology; D005260:Female; D006497:Heparan Sulfate / Q000032:analysis / Q000097:blood*; D006801:Humans; D008875:Middle Aged; D017698:Postmenopause / Q000097:blood*; D017697:Premenopause / Q000097:blood*; D011509:Proteoglycans / Q000032:analysis / Q000097:blood*; D013429:Sulfatases / Q000032:analysis / Q000097:blood*; D015238:Sulfotransferases / Q000032:analysis / Q000097:blood*
chemical_list: D006497:Heparan Sulfate; D011509:Proteoglycans; D013429:Sulfatases; D015238:Sulfotransferases
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