Amyloid fibril formation in vitro from halophilic metal binding protein: its high solubility and reversibility minimized formation of amorphous protein aggregations.
Amyloid fibril formation in vitro from halophilic metal binding protein: its high solubility and reversibility minimized formation of amorphous protein aggregations.
复制标题
嗜盐金属结合蛋白在体外形成淀粉样原纤维：其高溶解度和可逆性最大限度地减少了无定形蛋白聚集体的形成。
DOI：
10.1002/pro.2359
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发表时间：
2013
期刊：
影响因子：
8
通讯作者：
Sugimoto Y.
中科院分区：
文献类型：
--
作者：
Tokunaga Y;Matsumoto M;Tokunaga M;Arakawa T;Sugimoto Y.
嗜盐蛋白的特点是高净负电荷和相对较少的疏水氨基酸，使它们具有抗聚集性。本研究中使用的中等嗜盐菌salexigens嗜盐杆菌中富含组氨酸的金属结合蛋白（HP）也具有这些特性。在这里，我们研究了嗜盐蛋白如何在体外形成淀粉样纤维蛋白。通过硫黄素荧光、CD光谱和透射显微镜或原子力显微镜观察到，在pH 2.0和58°C下培养的HP很容易形成淀粉样蛋白原纤维。然而，在这些低pH的恶劣条件下，His - HP被迅速水解成更小的肽，最有可能负责淀粉样蛋白纤维的快速形成。从原纤维中分离出三种主要的酸水解肽，发现它们很容易形成原纤维。华尔兹软件预测在这些肽序列中形成原纤维的合成肽也形成了原纤维。当与10-20% 2,2,2 -三氟乙醇在pH 7.8和25°C下孵育时，没有肽键裂解，全长His - HP也很容易形成淀粉样蛋白纤维。
Halophilic proteins are characterized by high net negative charges and relatively small fraction of hydrophobic amino acids, rendering them aggregation resistant. These properties are also shared by histidine‐rich metal binding protein (HP) from moderate halophile,Chromohalobacter salexigens, used in this study. Here, we examined how halophilic proteins form amyloid fibrilsin vitro. His‐tagged HP, incubated at pH 2.0 and 58°C, readily formed amyloid fibrils, as observed by thioflavin fluorescence, CD spectra, and transmission or atomic force microscopies. Under these low‐pH harsh conditions, however, His‐HP was promptly hydrolyzed to smaller peptides most likely responsible for rapid formation of amyloid fibril. Three major acid‐hydrolyzed peptides were isolated from fibrils and turned out to readily form fibrils. The synthetic peptides predicted to form fibrils in these peptide sequences by Waltz software also formed fibrils. Amyloid fibril was also readily formed from full‐length His‐HP when incubated with 10–20% 2,2,2‐trifluoroethanol at pH 7.8 and 25°C without peptide bond cleavage.