Hydrolysis of a second Asp-Pro site at the N-terminus of NOTCH3 in inherited vascular dementia.
Hydrolysis of a second Asp-Pro site at the N-terminus of NOTCH3 in inherited vascular dementia.
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DOI：
10.1038/s41598-021-96679-9
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发表时间：
2021-08-26
影响因子：
4.6
通讯作者：
Wang MM
中科院分区：
文献类型：
--
作者：
Zhang X;Lee SJ;Wang MM
生化水平的脑血管病理学已通过伴有皮质下梗死和白质脑病的脑常染色体显性遗传性动脉病（CADASIL）的研究获得信息，CADASIL是一种由NOTCH 3突变引起的血管疾病。先前在CADASIL中的工作描述了通过蛋白质的第一个Asp-Pro序列的特异性非酶促切割产生的NOTCH 3的N-末端蛋白水解。在这里，我们研究了NOTCH 3的第二个Asp-Pro肽键（残基121-122）是否在CADASIL中被切割。产生单特异性抗体，其识别预测在Asp 121之后通过切割产生的新表位。这些抗体被用来定位切割事件在Asp 121在死后CADASIL和控制脑组织，并调查因素，调节切割Asp 121。我们报告说，在Asp 121裂解发生在高水平的CADASIL脑动脉的动脉介质。在白色物质中，软脑膜动脉比穿通动脉显示出更多的裂解产物，而对照血管仅含有少量裂解的NOTCH 3。蛋白水解在Asp 121发生在纯化的制备NOTCH 3胞外域，增加了酸性pH值和还原条件下，需要天然蛋白质构象裂解。增加NOTCH 3 EGF样结构域蛋白的浓度提高了蛋白水解的水平。另一方面，几种聚阴离子化学物质有效地阻断了Asp 121处的裂解。这些研究表明，CADASIL中的NOTCH 3蛋白在不稳定的Asp-Pro肽键处的多个位置被切割。因此，慢性脑血管疾病与其他神经退行性疾病一样，其特征在于病理蛋白质在多个位点的蛋白水解，这可能产生小的病理肽。
Cerebrovascular pathology at the biochemical level has been informed by the study of cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL), a vascular disorder caused by NOTCH3 mutations. Previous work in CADASIL described N-terminal proteolysis of NOTCH3 generated by specific non-enzymatic cleavage of the first Asp-Pro sequence of the protein. Here, we investigated whether the second Asp-Pro peptide bond (residues 121–122) of NOTCH3 is cleaved in CADASIL. Monospecific antibodies were generated that recognize the neo-epitope predicted to be generated by cleavage after Asp121. These antibodies were used to localize cleavage events at Asp121 in post-mortem CADASIL and control brain tissue and to investigate factors that regulate cleavage at Asp121. We report that cleavage at Asp121 occurs at a high level in the arterial media of CADASIL cerebral arteries. Leptomeningeal arteries demonstrated substantially more cleavage product than penetrating arteries in the white matter, and control vessels harbored only a small amount of cleaved NOTCH3. Proteolysis at Asp121 occurred in purified preparations of NOTCH3 ectodomain, was increased by acidic pH and reductive conditions, and required native protein conformation for cleavage. Increasing the concentration of NOTCH3 EGF-like domain protein elevated the level of proteolysis. On the other hand, several polyanionic chemicals potently blocked cleavage at Asp121. These studies demonstrate that the NOTCH3 protein in CADASIL is cleaved in multiple locations at labile Asp-Pro peptide bonds. As such, chronic brain vascular disease, like other neurodegenerative conditions, features proteolysis of pathological proteins at multiple sites which may generate small pathological peptides.
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影响因子：
2.9
作者：
Zhang, Xiaojie;Lee, Soo Jung;Young, Kelly Z.;Josephson, David A.;Geschwind, Michael D.;Wang, Michael M.
通讯作者：
Wang, Michael M.
影响因子：
2.1
作者：
Skribanek, Z;Mezo, G;Hudecz, F
通讯作者：
Hudecz, F
影响因子：
3.7
作者：
Cartee NMP;Lee SJ;Keep SG;Wang MM
通讯作者：
Wang MM
影响因子：
6.9
作者：
Gatti JR;Zhang X;Korcari E;Lee SJ;Greenstone N;Dean JG;Maripudi S;Wang MM
通讯作者：
Wang MM
影响因子：
4.8
作者：
Lidell, ME;Johansson, MEV;Hansson, GC
通讯作者：
Hansson, GC