The binding of chondroitin sulfate to pleiotrophin/heparin-binding growth-associated molecule is regulated by chain length and oversulfated structures
The binding of chondroitin sulfate to pleiotrophin/heparin-binding growth-associated molecule is regulated by chain length and oversulfated structures
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DOI：
10.1074/jbc.m507750200
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发表时间：
2006-02-24
影响因子：
4.8
通讯作者：
Hata, T
中科院分区：
文献类型：
--
作者：
Maeda, N;Fukazawa, N;Hata, T
Pleiotroin是一种18 kDa的肝素结合生长因子，以硫酸软骨素(CS)蛋白多糖、PTP Zeta为受体。有研究表明，CS中的D型结构(Glca(2S)β1-3GalNAc(6S))有助于PTP Zeta与多营养素的高亲和力结合。在这里，我们使用表面等离子体共振生物传感器分析鲨鱼软骨CS-D与多营养素的相互作用，以揭示D型结构的重要性。CS-D用软骨素酶ABC部分消化，用Superdex 75pg色谱柱分离。~gt；=18-聚CS组分与多营养素有明显的结合作用，且较长的组分对多营养素的亲和力强于较短的组分。接近46分子的CS组分以高亲和力(K-D=类似于30 nM)与密集固定的多核营养素结合，结合反应符合二价分析物模型。然而，当固定化多营养素的密度降低时，亲和力显著降低(K-D=2.5µM)，反应不再符合模型，被认为是单价结合。与24聚体相似的20个组分与密集固定的多聚体营养素(K-D=3，类似于20µM)表现出低亲和力结合，似乎是单价的。当用强阴离子交换高效液相色谱分离得到与22-聚壳低聚糖相似的低聚糖时，每个组分对多营养素的亲和力不同(K-D=0.36，类似于>；10muM)，并且亲和力与D-和E-(glca beta 1-3GalNAc(4S，6S))型过硫酸盐化结构的数量相关。这些结果表明，多营养素与CS的结合受CS的多价性和过量硫酸盐化结构的数量的调节，CS类似于20聚体。
Pleiotrophin is an 18-kDa heparin-binding growth factor, which uses chondroitin sulfate (CS) proteoglycan, PTP zeta as a receptor. It has been suggested that the D-type structure (GlcA(2S)beta 1-3GalNAc(6S)) in CS contributes to the high affinity binding between PTP zeta and pleiotrophin. Here, we analyzed the interaction of shark cartilage CS-D with pleiotrophin using a surface plasmon resonance biosensor to reveal the importance of D-type structure. CS-D was partially digested with chondroitinase ABC, and fractionated using a Superdex 75pg column. The >= 18-mer CS fractions showed significant binding to pleiotrophin, and the longer fractions had stronger affinity for pleiotrophin than the shorter ones. The similar to 46-mer CS fraction bound to densely immobilized pleiotrophin with high affinity (K-D = similar to 30 nM), and the binding reactions fitted the bivalent analyte model. However, when the density of the immobilized pleiotrophin was lowered, the strength of affinity remarkably decreased (K-D = similar to 2.5 mu M), and the reactions no longer fitted the model and were considered to be monovalent binding. The 20 similar to 24-mer fractions showed low affinity binding to densely immobilized pleiotrophin (K-D = 3 similar to 20 mu M), which seemed to be monovalent. When similar to 22-mer CS oligosaccharides were fractionated by strong anion exchange HPLC, each fraction differed in affinity for pleiotrophin (K-D = 0.36 similar to > 10 mu M), and the affinity correlated with the amounts of D- and E- (GlcA beta 1-3GalNAc(4S, 6S)) type oversulfated structures. These results suggest that the binding of pleiotrophin to CS is regulated by multivalency with CS similar to 20 mer as a unit and by the amounts of oversulfated structures.