Structural characterization of human liver heparan sulfate
Structural characterization of human liver heparan sulfate
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DOI：
10.1016/j.bbagen.2004.09.007
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发表时间：
2005-01-18
影响因子：
3
通讯作者：
Linhardt, RJ
中科院分区：
文献类型：
--
作者：
Vongchan, P;Warda, M;Linhardt, RJ
本文报道了人肝硫酸乙酰肝素的分离、纯化和结构鉴定。1H-1-NMR光谱证实了这种糖胺聚糖（GAG）的纯度，二维1H-1-NMR证实了它是硫酸乙酰肝素。酶解聚分离的硫酸乙酰肝素，然后通过梯度聚丙烯酰胺凝胶，证实了其肝素裂解酶的敏感性。使用具有柱后衍生化和荧光检测的反相离子对（RPIP）HPLC测定所得不饱和二糖的浓度。该分析的结果清楚地表明分离的GAG是硫酸乙酰肝素，而不是肝素。人肝硫酸乙酰肝素类似于肝素，因为它具有降低的未硫酸化二糖含量和升高的平均硫酸化水平。然而，人肝硫酸乙酰肝素的抗凝血酶介导的抗Xa因子活性远低于猪肠（药用）肝素，但与标准猪肠硫酸乙酰肝素相当。此外，人肝硫酸乙酰肝素显示出比从猪肝或从人肝癌Hep 2G细胞系分离的硫酸乙酰肝素更高的硫酸化程度。(C)2004 Elsevier B. V.保留所有权利。
The isolation, purification and structural characterization of human liver heparan sulfate are described. H-1-NMR spectroscopy demonstrates the purity of this glycosaminoglycan (GAG) and two-dimensional H-1-NMR confirmed that it was heparan sulfate. Enzymatic depolymerization of the isolated heparan sulfate, followed by gradient polyacrylamide gel, confirmed its heparin lyase sensitivity. The concentration of resulting unsaturated disaccharides was determined using reverse phase ion-pairing (RPIP) HPLC with post column derivatization and fluorescence detection. The results of this analysis clearly demonstrate that the isolated GAG was heparan sulfate, not heparin. Human liver heparan sulfate was similar to heparin in that it has a reduced content of unsulfated disaccharide and an elevated average sulfation level. The antithrombin-mediated anti-factor Xa activity of human liver heparan sulfate, however, was much lower than porcine intestinal (pharmaceutical) heparin but was comparable to standard porcine intestinal heparan sulfate. Moreover, human liver heparan sulfate shows higher degree of sulfation than heparan sulfate isolated from porcine liver or from the human hepatoma Hep 2G cell line. (C) 2004 Elsevier B.V. All rights reserved.