Heat-Induced Aggregation of Hen Ovalbumin Suggests a Key Factor Responsible for Serpin Polymerization.
Heat-Induced Aggregation of Hen Ovalbumin Suggests a Key Factor Responsible for Serpin Polymerization.
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DOI：
10.1021/acs.biochem.8b00619
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发表时间：
2018-08
期刊：
影响因子：
2.9
通讯作者：
Masahiro Noji;M. So;Kei-ichi Yamaguchi;H. Hojo;M. Onda;Yoko Akazawa-Ogawa;Y. Hagihara;Y. Goto
中科院分区：
文献类型：
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作者：
Masahiro Noji;M. So;Kei-ichi Yamaguchi;H. Hojo;M. Onda;Yoko Akazawa-Ogawa;Y. Hagihara;Y. Goto
卵清蛋白（OVA）是蛋清的主要成分，也是一种非抑制蛇形蛋白超家族蛋白，虽然有报道称其可形成纤维聚集体，但其与各种退行性疾病相关的淀粉样原纤维的关系尚不清楚。我们使用淀粉样蛋白特异性硫黄素T测定法或发光二极管灯直接成像以及几种物理化学方法研究了完整OVA的热诱导聚集，结果证实完整OVA形成聚集体，其中有一小部分淀粉样蛋白核心，主要是无定形聚集体。我们用胰蛋白酶水解分离淀粉样蛋白核心肽。分离的23个残基肽pOVA具有明显的淀粉样变性，对应于导致丝状蛋白病的丝状蛋白潜伏期转变和结构域交换聚合的一个关键区域（β-链3A）。虽然pOVA的强淀粉样变性在胰消化的混合物中被抑制，但在各种盐存在的酸性条件下观察到它，其中pOVA具有正电荷。细胞毒性测量表明，虽然热处理的OVA聚集体表现出最强的毒性，但这是由于无定形聚集体的一般性质，而不是淀粉样毒性。预测表明，β-链3A区域的高淀粉样变性是各种蛇蛋白的共同特征。这表明β-链3A的高淀粉样变性对丝氨酸蛋白潜伏期转移和结构域交换聚合很重要，在OVA中被保留，并在热聚集时形成β-棘淀粉样蛋白核。
Although ovalbumin (OVA), a main component of hen egg white and a non-inhibitory serpin superfamily protein, has been reported to form fibrillar aggregates, its relationship with amyloid fibrils associated with various degenerative diseases is unclear. We studied the heat-induced aggregation of intact OVA using an amyloid-specific thioflavin T assay with a fluorometer or direct imaging with a light-emitting diode lamp and several physicochemical approaches, and the results confirmed that intact OVA forms aggregates with a small part of amyloid cores and dominantly amorphous aggregates. We isolated the amyloidogenic core peptide by proteolysis with trypsin. The isolated 23-residue peptide, pOVA, with marked amyloidogenicity, corresponded to one (β-strand 3A) of the key regions involved in serpin latency transition and domain-swap polymerization leading to serpinopathies. Although the strong amyloidogenicity of pOVA was suppressed in a mixture of tryptic digests, it was observed under acidic conditions in the presence of various salts, with which pOVA has a positive charge. Cytotoxicity measurements suggested that, although heat-treated OVA aggregates exhibited the strongest toxicity, it was attributed to a general property of amorphous aggregates rather than amyloid toxicity. Predictions indicated that the high amyloidogenicity of the β-strand 3A region is common to various serpins. This suggests that the high amyloidogenicity of β-strand 3A that is important for serpin latency transition and domain-swap polymerization is retained in OVA and constitutes β-spine amyloid cores upon heat aggregation.