Expression of recombinant apopholasin using a baculovirus-silkworm multigene expression system and activation via dehydrocoelenterazine.,Bioorganic & Medicinal Chemistry Letters

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Expression of recombinant apopholasin using a baculovirus-silkworm multigene expression system and activation via dehydrocoelenterazine.
Bioorganic & Medicinal Chemistry Letters ( IF 2.5 ) Pub Date : 2020-04-08 , DOI: 10.1016/j.bmcl.2020.127177
Maiko Moriguchi ₁ , Ryo Takahashi ₂ , Bubwoong Kang ₁ , Masaki Kuse ₃

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Pholasin is a photoprotein derived from the glowing bivalve mollusk, Pholas dactylus. Even though the chemical structure of the prosthetic group (chromophore) responsible for the light emission character of the mollusk remains unknown, research has shown that the presence of dehydrocoelenterazine (DCL) increased light emission and that the dithiothreitol adduct of DCL was isolated from Pholasin®. To date, our research has been focused on activating apopholasin, the naturally occurring apoprotein of Pholasin®, using DCL. In the current study, the expression of recombinant apopholasin via a baculovirus-silkworm multigene expression system is reported. Additionally, the purification of apopholasin using a Flag®-affinity column, the activation of apopholasin using DCL, and the initiation of its luminescent character through the addition of a peroxidase-hydrogen peroxide mixture are reported. The peroxidase-H2O2-dependent luminescence was observed from the recombinant apopholasin activated with DCL.

中文翻译：

使用杆状病毒-蚕多基因表达系统表达重组载脂蛋白，并通过脱氢腔肠素激活。

荧光素是一种发光的双壳贝类软体动物，发光的Dactylus。尽管负责软体动物发光特性的辅基基团（生色团）的化学结构仍是未知的，但研究表明脱氢腔肠素（DCL）的存在增加了发光，并且DCL的二硫苏糖醇加合物是从Pholasin®中分离出来的。迄今为止，我们的研究集中在使用DCL激活自然产生的载脂蛋白Apopholasin。在当前的研究中，报道了通过杆状病毒-蚕多基因表达系统表达重组载脂蛋白。此外，使用Flag®-affinity柱纯化载脂蛋白，使用DCL活化载脂蛋白，报道了通过添加过氧化物酶-过氧化氢混合物引发其发光特性。从被DCL激活的重组apopholasin中观察到过氧化物酶-H2O2依赖性发光。

更新日期：2020-04-08

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