Demonstration of the pleiotrophin-binding oligosaccharide sequences isolated from chondroitin sulfate/dermatan sulfate hybrid chains of embryonic pig brains
Demonstration of the pleiotrophin-binding oligosaccharide sequences isolated from chondroitin sulfate/dermatan sulfate hybrid chains of embryonic pig brains
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DOI：
10.1074/jbc.m507304200
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发表时间：
2005-10-21
影响因子：
4.8
通讯作者：
Sugahara, K
中科院分区：
文献类型：
--
作者：
Bao, XF;Muramatsu, T;Sugahara, K
哺乳动物脑中含有大量的硫酸软骨素（CS）、硫酸皮肤素（DS）和CS/ DS杂合链。从胚胎猪脑中分离的CS/ DS链（E-CS/ DS）通过与多效生长因子（PTN）（一种肝素结合生长因子）相互作用促进培养的胚胎小鼠海马神经元中神经突的生长。在这里，我们分析了从E-CS/ DS中分离的寡糖，这表明八糖是能够在生理盐浓度下与PTN相互作用的最小尺寸。通过酶消化，然后进行PTN亲和层析，分别从荧光标记的PTN结合和未结合的八糖组分中纯化了五个和八个序列。它们的序列通过酶消化结合高效液相色谱法测定，揭示了过硫酸化D和/或iD二糖在对PTN的低但显著的亲和力中的关键作用，PTN是轴突发生所需的。关键的D和iD单元分别是GlcUA（2- O-硫酸酯）β 1 - 3GalNAc（6- O-硫酸酯）和IdoUA（2- O-硫酸酯）α 1 - 3GalNAc（6- O-硫酸酯），其中IdoUA代表L-艾杜糖醛酸.相反，与PTN的高亲和力相互作用需要具有E单元的十糖（GlcUA β 1 - 3GalNAc（4，6- O-二硫酸盐）），B单位（GlcUA（2- O-硫酸盐）β 1 - 3GalNAc（4- O-硫酸盐））和/或它们的含IdoUA的对应物（iE和iB）除了D/ iD单位，虽然这种强相互作用的生物学意义仍有待研究。因此，链大小和组成对于与PTN的相互作用至关重要，并且PTN以不同的亲和力结合E-CS/ DS链中的多个序列。值得注意的是，不仅硫酸乙酰肝素，而且哺乳动物大脑的CS/ DS混合链结构含有高度的微异质性与一簇过硫酸化二糖，似乎在调节PTN的功能中发挥作用。
Mammalian brains contain significant amounts of chondroitin sulfate ( CS), dermatan sulfate ( DS), and CS/ DS hybrid chains. CS/ DS chains isolated from embryonic pig brains ( E- CS/ DS) promote the outgrowth of neurites in embryonic mouse hippocampal neurons in culture by interacting with pleiotrophin ( PTN), a heparin-binding growth factor. Here, we analyzed oligosaccharides isolated from E- CS/ DS, which showed that octasaccharides were the minimal size capable of interacting with PTN at a physiological salt concentration. Five and eight sequences were purified from fluorescently labeled PTN- bound and - unbound octasaccharide fractions, respectively, by enzymatic digestion followed by PTN- affinity chromatography. Their sequences were determined by enzymatic digestion in conjunction with high performance liquid chromatography, revealing a critical role for oversulfated D and/ or iD disaccharides in the low yet significant affinity for PTN, which is required for neuritogenesis. The critical D and iD units are GlcUA( 2- O- sulfate) beta 1 - 3GalNAc( 6- O- sulfate) and IdoUA( 2- O- sulfate) alpha 1 - 3GalNAc( 6- O- sulfate), respectively, where IdoUA represents L- iduronic acid. In contrast, high affinity interactions with PTN required decasaccharides with E units ( GlcUA beta 1 - 3GalNAc( 4, 6- O- disulfate)), B units ( GlcUA( 2- O- sulfate) beta 1 - 3GalNAc( 4- O- sulfate)), and/ or their IdoUA- containing counterparts ( iE and iB) in addition to D/ iD units, although the biological significance of such strong interactions remains to be investigated. Thus, chain size and composition are crucial to the interaction with PTN, and PTN binds to multiple sequences in E- CS/ DS chains with distinct affinity. Notably, not only heparan sulfate but also CS/ DS hybrid chain structures of mammalian brains contain a high degree of microheterogeneity with a cluster of oversulfated disaccharides and appear to play roles in regulating the functions of PTN.