Division of Chemical Biology and Medicinal Chemistry, Eshelman School of Pharmacy, University of North Carolina, Chapel Hill, North Carolina 27599, USA.
J Biol Chem. 2012 Aug 17;287(34):29054-61. doi: 10.1074/jbc.M112.358523. Epub 2012 Jul 6.
Heparan sulfate (HS) and heparin are highly sulfated polysaccharides. Heparin is a commonly used anticoagulant drug that inhibits the activities of factors Xa and IIa (also known as thrombin) to prevent blood clot formation. Here, we report the synthesis of a series of size-defined oligosaccharides to probe the minimum size requirement for an oligosaccharide with anti-IIa activity. The synthesis was completed by a chemoenzymatic approach involving glycosyltransferases, HS sulfotransferases, and C(5)-epimerase. We demonstrate the ability to synthesize highly purified N-sulfo-oligosaccharides having up to 21 saccharide residues. The results from anti-Xa and anti-IIa activity measurements revealed that an oligosaccharide longer than 19 saccharide residues is necessary to display anti-IIa activity. The oligosaccharides also exhibit low binding toward platelet factor 4, raising the possibility of preparing a synthetic heparin with a reduced effect of heparin-induced thrombocytopenia. The results from this study demonstrate the ability to synthesize large HS oligosaccharides and provide a unique tool to probe the structure and function relationships of HS that require the use of large HS fragments.
硫酸乙酰肝素 (HS) 和肝素是高度硫酸化的多糖。肝素是一种常用的抗凝药物，通过抑制因子 Xa 和 IIa（也称为凝血酶）的活性来防止血栓形成。在这里，我们报告了一系列大小确定的寡糖的合成，以探究具有抗 IIa 活性的寡糖的最小尺寸要求。该合成是通过涉及糖基转移酶、HS 硫酸转移酶和 C(5)-差向异构酶的化学酶法完成的。我们证明了能够合成具有多达 21 个糖残基的高度纯化的 N-磺酸寡糖。抗 Xa 和抗 IIa 活性测量的结果表明，需要具有 19 个以上糖残基的寡糖才能显示抗 IIa 活性。这些寡糖对血小板因子 4 的结合也较低，这提高了制备具有降低肝素诱导血小板减少症作用的合成肝素的可能性。这项研究的结果证明了合成大型 HS 寡糖的能力，并提供了一种独特的工具来研究需要使用大型 HS 片段的 HS 的结构和功能关系。
