Organ-specific Heparan Sulfate Structural Phenotypes

Abstract

The functions of heparan sulfate (HS) depend on the expression of structural domains that interact with protein partners. Glycosaminoglycans (GAGs) exhibit a high degree of polydispersity in their composition, chain length, sulfation, acetylation, and epimerization patterns. It is essential for the understanding of GAG biochemistry to produce detailed structural information as a function of spatial and temporal factors in biological systems. Toward this end, we developed a set of procedures to extract GAGs from various rat organ tissues and examined and compared HS expression levels using liquid chromatography/ mass spectrometry. Here we demonstrate detailed variations in HS GAG chains as a function of organ location. These studies shed new light on the structural variation of GAG chains with respect to average length, disaccharide composition, and expression of low abundance structural epitopes, including unsubstituted amino groups and lyase-resistant oligosaccharides. The data show the presence of a disaccharide with an unsubstituted amino group that is endogenous and widely expressed in mammalian organ tissues.

Extended N-Sulfated Domains Reside at the Nonreducing End of Heparan Sulfate Chains

“…In addition to this structure, the isomer pair U0S6/ U2S0 was enriched. This observation is similar to our previous study of HS disaccharides from rat organs (23), and it may represent an evolutionary similarity between these two organisms.…”

“…Disaccharide Analysis Using Size-exclusion LC/MS-Disaccharides produced by exhaustive lyase digestion were analyzed directly after depolymerization using size-exclusion chromatography (SEC)-mass spectrometry as described previously (23). Briefly, HS samples (1 g of material) were injected onto a Superdex TM peptide PC 3.2/30 column (GE Healthcare) on line with an ABI SCIEX Q-Star Pulsar TOF mass spectrometer operating in the negative-ion mode.…”

“…By comparing the saturated and ⌬-un-saturated disaccharide profiles, the probability that a given disaccharide will appear at the NRE compared with the internal region of the chain can be assessed (Fig. 1B) (23). Strikingly, all five of the organ HS preparations exhibit enrichment of the structure U0S0 at the NRE.…”

Organ-specific Sulfation Patterns of Heparan Sulfate Generated by Extracellular Sulfatases Sulf1 and Sulf2 in Mice

“…The percentages were relatively high (Ͼ10%) for ⌬UA2S-GlcNS6S in the liver (16.7%) and kidney (10.7%), for ⌬UA-GlcNS6S in the spleen (14.9%) and kidney (10.7%), for ⌬UA2S-GlcNS in the lung (17.2%), brain (14.7%), and testis (13.7%), and for ⌬UA-GlcNAc6S in the spleen (12.6%), kidney (12.2%), and liver (10.1%). These sulfation profiles of HS concur well with those previously reported in mouse and bovine organs (37)(38)(39)(40).…”

Remodeling of Marrow Hematopoietic Stem and Progenitor Cells by Non-self ST6Gal-1 Sialyltransferase

“…In some instances, the released oligosaccharides were further enriched into acidic, sialic acid-containing, N-linked glycans using a modified procedure based on a glycosaminoglycan purification protocol described previously by Shi and Zaia (18). A macrospin column (Harvard Apparatus) with DEAE resins was saturated by washing with 300 l of elution buffer (2 M NH 4 HCO 3 ) and then 3 ϫ 300 l of loading buffer (5 mM TrisHCl, pH 8).…”