Comparative equilibrium denaturation studies of the neurotrophins: nerve growth factor, brain-derived neurotrophic factor, neurotrophin 3, and neurotrophin 4/5.
Comparative equilibrium denaturation studies of the neurotrophins: nerve growth factor, brain-derived neurotrophic factor, neurotrophin 3, and neurotrophin 4/5.
复制标题
神经营养蛋白的平衡变性比较研究：神经生长因子、脑源性神经营养因子、神经营养蛋白 3 和神经营养蛋白 4/5。
DOI：
10.1021/bi00181a602
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发表时间：
1994
期刊：
影响因子：
2.9
通讯作者：
Neet,KE
中科院分区：
文献类型：
--
作者：
Timm,DE;deHaseth,PL;Neet,KE
修订稿于 1994 年 2 月 8 日收到® 摘要：神经营养蛋白是脊椎动物神经元发育和存活所需的小二聚体蛋白家族。使用溶剂变性研究来比较重组人神经生长因子 (hNGF)、脑源性神经营养因子 (BDNF)、神经营养蛋白 3 (NT-3) 和神经营养蛋白 4/5 (NT-4/5) 与从小鼠颌下腺分离的神经生长因子 (mNGF)。尽管该家族之间保留了超过 50% 的序列同一性，但这些蛋白质的折叠和解折叠揭示了显着的结构差异。通过荧光强度、荧光偏振和圆二色性监测盐酸胍的变性和 pH 7 和 3.5 下的复性。所有四种神经营养素的平衡展开曲线的中点与技术无关，但取决于蛋白质浓度，表明涉及天然神经营养素二聚体和变性神经营养素单体（N2=2D）的双态模型描述了折叠和未折叠神经营养素之间的平衡。二聚体神经营养素的构象稳定性表明，mNGF 的构象稳定性最低（19.3 kcal/mol）； hNGF、NT-3和NT-4/5具有中等稳定性，BDNF具有最高稳定性（26.4 kcal/mol）。去除变性剂后天然光谱特征的恢复表明解折叠过程是可逆的。因此，NGF或NT-4/5在pH 7和3.5下以及BDNF在pH 3.5下的解折叠和重折叠曲线是一致的。然而，BDNF 和 NT-3 的解折叠和重折叠曲线在 pH 7 时并不重合。神经营养素的稳定性随着 pH 的降低而降低，并且具有紧凑的单体中间体 (N2=[21]=
Revised Manuscript Received February 8, 1994® abstract: The neurotrophins are a family of small dimeric proteins required for the development and survival of vertebrate neurons. Solvent denaturation studies were used tocompare recombinant human nerve growth factor (hNGF), brain-derived neurotrophic factor (BDNF), neurotrophin 3 (NT-3), and neurotrophin 4/5 (NT-4/5) to nerve growth factor isolated from mouse submaxillary glands (mNGF). Although greater than 50% sequence identity is conserved among this family, significant structural differences were revealed by the folding and unfolding of these proteins. Denaturation in guanidine hydrochloride and renaturation at pH 7 and 3.5 were monitored by fluorescence intensity, fluorescence polarization, and circular dichroism. The midpoint of equilibrium unfolding curves for all four neurotrophins was independent of the technique but was dependent on protein concentration, indicating that a two-state model involving native neurotrophin dimers and denatured neurotrophin monomers(N2=2D) describes the equilibrium between folded and unfolded neurotrophins. The conformational stabilities of the dimeric neurotrophins revealed that mNGF had the lowest conformational stability (19.3 kcal/mol); hNGF, NT-3, and NT-4/5 had intermediate stabilities, and BDNF had the highest stability (26.4 kcal/mol). Recovery of native spectroscopic characteristics upon removal of denaturant indicated that the unfolding process is reversible. Accordingly, unfolding and refolding curves were coincident formNGF or NT-4/5 at pH 7 and 3.5 and for BDNF at pH 3.5. However, BDNF and NT-3 unfolding and refolding curves were not coincident at pH 7. The stability of the neurotrophins decreased as pH decreased, with compact monomeric intermediates (N2=[21]=