Institute of Integrative Biology, Department of Chemical and Structural Biology, University of Liverpool, Crown Street, Liverpool L69 7ZB, United Kingdom.
J Biol Chem. 2012 Nov 16;287(47):40061-73. doi: 10.1074/jbc.M112.398826. Epub 2012 Sep 27.
The functions of a large number (>435) of extracellular regulatory proteins are controlled by their interactions with heparan sulfate (HS). In the case of fibroblast growth factors (FGFs), HS binding determines their transport between cells and is required for the assembly of high affinity signaling complexes with their cognate FGF receptor. However, the specificity of the interaction of FGFs with HS is still debated. Here, we use a panel of FGFs (FGF-1, FGF-2, FGF-7, FGF-9, FGF-18, and FGF-21) spanning five FGF subfamilies to probe their specificities for HS at different levels as follows: binding parameters, identification of heparin-binding sites (HBSs) in the FGFs, changes in their secondary structure caused by heparin binding and structures in the sugar required for binding. For interaction with heparin, the FGFs exhibit K(D) values varying between 38 nM (FGF-18) and 620 nM (FGF-9) and association rate constants spanning over 20-fold (FGF-1, 2,900,000 M(-1) s(-1) and FGF-9, 130,000 M(-1) s(-1)). The canonical HBS in FGF-1, FGF-2, FGF-7, FGF-9, and FGF-18 differs in its size, and these FGFs have a different complement of secondary HBS, ranging from none (FGF-9) to two (FGF-1). Differential scanning fluorimetry identified clear preferences in these FGFs for distinct structural features in the polysaccharide. These data suggest that the differences in heparin-binding sites in both the protein and the sugar are greatest between subfamilies and may be more restricted within a FGF subfamily in accord with the known conservation of function within FGF subfamilies.
大量（>435）细胞外调节蛋白的功能受其与硫酸乙酰肝素（HS）相互作用的控制。对于成纤维细胞生长因子（FGFs），HS 结合决定了它们在细胞之间的运输，并与它们同源的 FGF 受体形成高亲和力信号复合物是必需的。然而，FGFs 与 HS 的相互作用特异性仍存在争议。在这里，我们使用一组涵盖五个 FGF 亚家族的 FGFs（FGF-1、FGF-2、FGF-7、FGF-9、FGF-18 和 FGF-21）来从以下几个方面探测它们与 HS 的特异性：结合参数、鉴定 FGF 中的肝素结合位点（HBSs）、肝素结合引起的二级结构变化以及结合所需的糖的结构。对于与肝素的相互作用，FGFs 的 K(D) 值在 38 nM（FGF-18）和 620 nM（FGF-9）之间变化，而结合速率常数跨越 20 倍（FGF-1、2、900、000 M(-1) s(-1) 和 FGF-9、130、000 M(-1) s(-1)）。FGF-1、FGF-2、FGF-7、FGF-9 和 FGF-18 中的典型 HBS 在大小上有所不同，并且这些 FGFs 具有不同的二级 HBS 互补，从无（FGF-9）到两个（FGF-1）。差示扫描荧光法确定了这些 FGFs 在多糖中对不同结构特征的明显偏好。这些数据表明，在蛋白质和糖中肝素结合位点的差异在亚家族之间最大，并且在 FGF 亚家族内可能更受限制，这与 FGF 亚家族内功能的已知保守性一致。
