Usui Masakatsu, Harada Akihito, Ishimaru Takayuki, Sakumichi Emiri, Saratani Fumihiko, Sato-Minami Chiho, Azakami Hiroyuki, Miyasaki Taiko, Hanaoka Ken'ichi
Department of Food Science and Technology, National Fisheries University, Nagata-Honmachi, Shimonoseki, Japan.
Biosci Biotechnol Biochem. 2013;77(5):948-53. doi: 10.1271/bbb.120887. Epub 2013 May 7.
Tropomyosins are common heat-stable crustacean allergens. However, their heat stability and their effects on antigenicity have not been clarified. We purified tropomyosin in this study from raw kuruma prawns (Marsupenaeus japonicus) without heat processing. SDS-PAGE of the purified protein showed a band at approximately 35 kDa that cross-reacted with IgE from the serum of a shrimp-allergic patient, identifying it as Pen j 1. The circular dichroism spectrum of native Pen j 1 revealed the common α-helical structure of tropomyosins which easily collapsed upon heating to 80 °C. However, there were no insoluble aggregates after heating, and the protein regained its native CD spectral pattern after cooling to 25 °C. There was no significant difference in total IgG production between mice sensitized with native and heated Pen j 1. These results suggest that heat-denatured Pen j 1 refolded upon cooling and maintained its antigenicity following the heat treatment.
原肌球蛋白是常见的热稳定甲壳类过敏原。然而，它们的热稳定性及其对抗原性的影响尚未阐明。在本研究中，我们从未经热处理的生日本对虾（日本囊对虾）中纯化了原肌球蛋白。纯化蛋白的SDS-PAGE显示在约35 kDa处有一条带，与虾过敏患者血清中的IgE发生交叉反应，将其鉴定为Pen j 1。天然Pen j 1的圆二色光谱揭示了原肌球蛋白常见的α-螺旋结构，该结构在加热至80°C时容易塌陷。然而，加热后没有不溶性聚集体，冷却至25°C后蛋白质恢复了其天然CD光谱模式。用天然和加热的Pen j 1致敏的小鼠之间总IgG产生没有显著差异。这些结果表明，热变性的Pen j 1在冷却后重新折叠，并在热处理后保持其抗原性。
