Structural basis and potential role of heparin/heparan sulfate binding to the angiogenesis inhibitor endostatin
Structural basis and potential role of heparin/heparan sulfate binding to the angiogenesis inhibitor endostatin
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DOI：
10.1093/emboj/18.22.6240
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发表时间：
1999-11-15
期刊：
影响因子：
11.4
通讯作者：
Timpl, R
中科院分区：
文献类型：
--
作者：
Sasaki, T;Larsson, H;Timpl, R
由哺乳动物细胞产生的重组小鼠内皮抑素与肝素结合的 Kd 为 0.3 μM，表明这种相互作用可能在其抗血管生成活性中发挥作用。丙氨酸诱变证明四个簇状精氨酸的主要位点（位置 155、158、184 和 270）和第二个位点（R193、R194）对于结合至关重要。相同的表位还参与内皮抑素与硫酸乙酰肝素和硫苷脂的结合，但不参与其与细胞外蛋白配体 fibulin-1 和 fibulin-2 的结合，对各种肝素片段的分析表明，有效结合内皮抑素的最小尺寸（12mer）以及 2-O- 和 6-O-硫酸化的关键作用，此外，从各种组织获得的硫酸乙酰肝素链的大部分（10-50%）显示出独特的与内皮抑素结合，表明其与细胞外和/或膜结合蛋白聚糖相互作用的潜力。在鸡绒毛尿囊膜测定中，由碱性成纤维细胞生长因子-2（FGF-2）而非血管内皮生长因子（VEGF）诱导的血管生成可以被内皮抑素以剂量依赖性方式抑制。肝素结合的突变阻断将内皮抑素抑制降低至低水平，但消除锌结合没有效果。
Recombinant mouse endostatin produced by mammalian cells was shown to bind to heparin with a Kd of 0.3 mu M, suggesting that this interaction may play a role in its anti-angiogenic activity. Alanine mutagenesis demonstrated that a major site of four clustered arginines (positions 155, 158, 184 and 270) and a second site (R193,R194) are essential for binding. The same epitopes also participate in endostatin binding to heparan sulfate and sulfatides but not in its binding to the extracellular protein ligands fibulin-1 and fibulin-2, Analyses with various heparin fragments demonstrated a minimum size (12mer) for efficient binding to endostatin and a crucial role of 2-O- and 6-O-sulfation, Furthermore, a substantial proportion (10-50%) of heparan sulfate chains obtained from various tissues showed a distinct binding to endostatin, indicating its potential to interact with extracellular and/or membrane-bound proteoglycans, Angiogenesis induced by basic fibroblast growth factor-2 (FGF-2), but not by vascular endothelial growth factor (VEGF), in a chick chorioallantoic membrane assay could be inhibited by endostatin in a dose-dependent manner. The mutational block of heparin binding decreased endostatin inhibition to low levels but elimination of zinc binding had no effect.