The major determinant of the heparin binding of glial cell-line-derived neurotrophic factor is near the N-terminus and is dispensable for receptor binding
The major determinant of the heparin binding of glial cell-line-derived neurotrophic factor is near the N-terminus and is dispensable for receptor binding
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DOI：
10.1042/bj20061747
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发表时间：
2007-05-15
影响因子：
4.1
通讯作者：
Rider, Christopher C.
中科院分区：
文献类型：
--
作者：
Alfano, Ivan;Vora, Parvez;Rider, Christopher C.
胶质细胞源性神经营养因子(GDNF)以及与肝素密切相关的细胞因子青蒿素和神经图灵与肝素结合强烈。GDNF的转化生长因子结构域的第一个半胱氨酸的16个氨基酸N端的富含碱性氨基酸的序列的缺失导致肝素结合显著减少，而邻近序列的去除和用丙氨酸取代其他碱性残基对没有影响。肝素结合序列与高亲和力GDNF多肽受体GFRα1(GDNF家族受体α1)的结合部位完全不同，肝素结合的GDNF能够同时与GFRα1结合。GDNF的肝素结合序列对于GFRα1结合和体外轴突生长实验都是必不可少的。令人惊讶的是，在本实验中观察到的野生型蛋白对GDNF生物活性的抑制仍然存在，缺失突变体缺乏肝素结合序列。肝素既不抑制也不增强GDNF-GFRα1的相互作用，GFRα1的胞外区不与肝素本身结合，排除了肝素交叉连接细胞因子和受体多肽的作用。肝素和硫酸肝素在GDNF信号转导中的作用尚不清楚，但本研究表明，它不发生在该途径的第一步，即GDNF-GFRα1结合。
GDNF (glial cell-line-derived neurotrophic factor), and the closely related cytokines artemin and neurturin, bind strongly to heparin. Deletion of a basic amino-acid-rich sequence of 16 residues N-terminal to the first cysteine of the transforming growth factor domain of GDNF results in a marked reduction in heparin binding, whereas removal of a neighbouring sequence, and replacement of pairs of other basic residues with alanine had no effect. The heparin-binding sequence is quite distinct from the binding site for the high affinity GDNF polypeptide receptor, GFR alpha 1 (GDNF family receptor alpha 1), and heparin-bound GDNF is able to bind GFR alpha 1 simultaneously. The heparin-binding sequence of GDNF is dispensable both for GFR alpha 1 binding, and for activity for in vitro neurite outgrowth assay. Surprisingly, the observed inhibition of GDNF bioactivity with the wild-type protein in this assay was still found with the deletion mutant lacking the heparin-binding sequence. Heparin neither inhibits nor potentiates GDNF-GFR alpha 1 interaction, and the extracellular domain of GFR alpha 1 does not bind to heparin itself, precluding heparin cross-bridging of cytokine and receptor polypeptides. The role of heparin and heparan sulfate in GDNF signalling remains unclear, but the present study indicates that it does not occur in the first step of the pathway, namely GDNF-GFR alpha 1 engagement.