Glycobiology Advance Access originally published online on April 20, 2005
Glycobiology 2005 15(8):818-826; doi:10.1093/glycob/cwi064
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Compositional profiling of heparin/heparan sulfate using mass spectrometry: assay for specificity of a novel extracellular human endosulfatase
3 Department of Chemistry, University of California, Berkeley, CA 94720; 4 Department of Anatomy, University of California, San Francisco, CA 94143; 5 UCSF Comprehensive Cancer Center, University of California, San Francisco, CA 94143; 6 Department of Molecular and Cell Biology, University of California, Berkeley, CA 94720; 7 Howard Hughes Medical Institute, University of California, Berkeley, CA 94720
1 To whom correspondence should be addressed; e-mail: jaleary{at}ucdavis.edu
2 Present address: Department of Chemistry and Division of Molecular and Cellular Biology, Genome Center, University of California Davis, One Shields Road, Davis, CA 95616
Received on February 4, 2005; revised on April 13, 2005; accepted on April 14, 2005
An important class of carbohydrates studied within the field of glycobiology, heparin and heparan sulfate (HS) have been implicated in a diverse array of biological functions. Changes in their sulfation pattern and domain organization have been associated with different pathological situations such as viral infectivity, tumor growth, and metastasis. To obtain structural information about these biomolecules, and the modifications they may undergo during different stages of cell growth and development, a mass spectrometry-based method was developed and used to obtain unambiguous structural information on the glycosaminoglycans (GAGs) that comprise heparin/HS. The method was applied to assay for the heparin substrate specificity of a newly discovered human extracellular endosulfatase, HSulf-2, which has been implicated in tumorigenesis. This new protocol incorporates 12 known heparin disaccharides, including three sets of isomers. A unique response factor (R) is determined for each disaccharide, whereas a multiplexed and data processing method is incorporated for faster data acquisition and quantification purposes. Proof of principle was performed by using various heparin/HS samples isolated from bovine and porcine tissues.
Key words: glycosaminoglycans / heparan sulfate / heparin / mass spectrometry / sulfatase assay
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