Neurotrophic activity of neudesin, a novel extracellular heme-binding protein, is dependent on the binding of heme to its cytochrome b5-like heme/steroid-binding domain
Neurotrophic activity of neudesin, a novel extracellular heme-binding protein, is dependent on the binding of heme to its cytochrome b5-like heme/steroid-binding domain
复制标题
DOI：
10.1074/jbc.m706679200
复制
发表时间：
2008-02-15
影响因子：
4.8
通讯作者：
Fujimoto, Masafumi
中科院分区：
文献类型：
--
作者：
Kimura, Ikuo;Nakayama, Yoshiaki;Fujimoto, Masafumi
Neudesin是神经元和未分化神经细胞中具有神经营养活性的分泌蛋白。我们在这里报告neudesin是一种细胞外血红素结合蛋白，其神经营养活性依赖于血红素与其细胞色素B（5）样血红素/类固醇结合结构域的结合。首先，我们发现至少一部分纯化的重组neudesin似乎与氯化血红素结合，因为纯化的neudesin溶液被染成绿色并在402 nm处具有尖锐的吸收峰。此外，外源性氯化血红素广泛增加氯化血红素结合neudesin的量。相比之下，neudesin三角洲HBD，缺乏血红素结合域的突变体，不能结合氯化血红素。重组neudesin结合外源性氯化血红素（neudesin-hemin）的神经营养活性显着大于重组neudesin在原代培养的神经元或Neuro 2a细胞，表明neudesin的活性依赖于氯化血红素。neudesin的神经营养活性的Fe（III）-原卟啉IX的结合增强，但无论是Fe（II）-原卟啉IX，也不是原卟啉IX单独。通过RNA干扰抑制内源性neudesin显著降低Neuro 2a细胞中的细胞存活。这表明内源性neudesin可能含有氯化血红素。用抗neudesin抗体进行的实验表明，在Neuro 2a细胞的培养基中检测到的内源性neudesin与氯化血红素相关，因为它根本不保留在血红素亲和柱上。Neudesin是第一个细胞外血红素结合蛋白，显示信号转导活动本身。目前的发现可能为细胞外血红素结合蛋白的功能提供新的线索。
Neudesin is a secreted protein with neurotrophic activity in neurons and undifferentiated neural cells. We report here that neudesin is an extracellular heme-binding protein and that its neurotrophic activity is dependent on the binding of heme to its cytochrome b(5)-like heme/steroid-binding domain. At first, we found that at least a portion of the purified recombinant neudesin appeared to bind hemin because the purified neudesin solution was tinged with green and had a sharp absorbance peak at 402 nm. The addition of exogenous hemin extensively increased the amount of hemin-bound neudesin. In contrast, neudesin Delta HBD, a mutant lacking the heme-binding domain, could not bind hemin. The neurotrophic activity of the recombinant neudesin that bound exogenous hemin (neudesin-hemin) was significantly greater than that of the recombinant neudesin in either primary cultured neurons or Neuro2a cells, suggesting that the activity of neudesin depends on hemin. The neurotrophic activity of neudesin was enhanced by the binding of Fe(III)-protoporphyrin IX, but neither Fe(II)-protoporphyrin IX nor protoporphyrin IX alone. The inhibition of endogenous neudesin by RNA interference significantly decreased cell survival in Neuro2a cells. This indicates that endogenous neudesin possibly contains hemin. The experiment with anti-neudesin antibody suggested that the endogenous neudesin detected in the culture medium of Neuro2a cells was associated with hemin because it was not retained on a heme-affinity column at all. Neudesin is the first extracellular heme-binding protein that shows signal transducing activity by itself. The present findings may shed new light on the function of extracellular heme-binding proteins.