The Binding Properties of Minimal Oligosaccharides Reveal a Common Heparan Sulfate/Dermatan Sulfate-binding Site in Hepatocyte Growth Factor/Scatter Factor That Can Accommodate a Wide Variety of Sulfation Patterns
The Binding Properties of Minimal Oligosaccharides Reveal a Common Heparan Sulfate/Dermatan Sulfate-binding Site in Hepatocyte Growth Factor/Scatter Factor That Can Accommodate a Wide Variety of Sulfation Patterns
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DOI：
10.1074/jbc.m807671200
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发表时间：
2009-03-06
影响因子：
4.8
通讯作者：
Lyon, Malcolm
中科院分区：
文献类型：
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作者：
Deakin, Jon A.;Blaum, Baerbel S.;Lyon, Malcolm
硫酸乙酰肝素 (HS)/肝素和硫酸皮肤素 (DS) 均以高亲和力与肝细胞生长因子/分散因子 (HGF/SF) 结合，并在体外作为必要的辅助因子发挥作用。这两种结构不同的糖胺聚糖（GAG）是如何被识别的仍不清楚。现在，我们使用一组可变但明确的硫酸化模式的最小三糖和四糖序列，结合凝胶迁移率变化测定的进一步开发，解决了这个问题，以允许同时比较不同寡糖的相对蛋白质亲和力/选择性。从这种方法看来，最小的结合序列是由侧翼为单硫酸化己糖胺残基的内部艾杜糖醛酸组成的二硫酸化三糖，并且额外的硫酸化进一步增强了亲和力。然而，HS/肝素和 DS 识别上的相似性似乎主要是由于缺乏硫酸化的任何明显位置要求。因此，仅含有两个硫酸盐的 HS/肝素四糖异构体，无论它们是纯 N-、2-O-还是 6-O-硫酸盐，都以与二硫酸化 DS 四糖相同的表观亲和力结合。此外，通过用肝素或 DS 寡糖滴定在 NK1（由 N 末端和第一个 Kringle 结构域组成的 HGF/SF 的截短变体）中诱导的 NMR 化学位移强烈表明两者结合到基本相同的位点。总之，这些观察结果揭示了 GAG-HGF/SF 界面中意想不到的灵活性，允许蛋白质中的单个结合位点适应来自不同 GAG 的可变硫酸化模式和/或密度的含艾杜糖醛酸序列。
Heparan sulfate (HS)/heparin and dermatan sulfate (DS) both bind with high affinity to hepatocyte growth factor/scatter factor (HGF/SF) and function as necessary co-factors in vitro. How both these two structurally distinct glycosaminoglycans (GAGs) are recognized has remained unclear. We have now reconciled this issue using a panel of minimal tri- and tetrasaccharide sequences of variable but well defined sulfation patterns in combination with further development of the gel mobility shift assay to allow simultaneous comparisons of relative protein affinities/selectivities for different oligosaccharides. From this approach it would seem that a minimum binding sequence is a disulfated trisaccharide comprised of an internal iduronate flanked by monosulfated hexosamine residues and that additional sulfation further enhances affinity. However, the similarity in recognition of HS/heparin and DS seems to arise primarily from a lack of any apparent positional requirement for sulfation. Thus, isomers of HS/heparin tetrasaccharides containing only two sulfates irrespective of whether they are purely N-, 2-O-, or 6-O-sulfates bind with equivalent apparent affinity as a disulfated DS tetrasaccharide. In addition, the NMR chemical shifts induced in NK1 (the truncated variant of HGF/SF comprised of the N- terminal and first Kringle domains) by titration with either heparin or DS oligosaccharides strongly indicate that both bind to essentially the same site. Together, these observations reveal an unexpected degree of flexibility in the GAG-HGF/SF interface, allowing a single binding site in the protein to accommodate iduronate-containing sequences of variable sulfation pattern and/or density from different GAGs.