Nishiguchi Tomoki, Yamada Toshimichi, Nasu Yusuke, Ito Mashiho, Yoshimura Hideaki, Ozawa Takeaki
J Biomed Opt. 2015 Oct;20(10):101205. doi: 10.1117/1.JBO.20.10.101205.
Luciferase, a bioluminescent protein, has been used as an analytical tool to visualize intracellular phenomena. Luciferase with red light emission is particularly useful for bioluminescence imaging because of its high transmittance in mammalian tissues. However, the luminescence intensity of existing luciferases with their emission over 600 nm is insufficient for imaging studies because of their weak intensities. We developed mutants of Emerald luciferase (Eluc) from Brazilian click beetle (Pyrearinus termitilluminans), which emits the strongest bioluminescence among beetle luciferases. We successively introduced four amino acid mutations into the luciferase based on a predicted structure of Eluc using homology modeling. Results showed that quadruple mutations R214K/H241K/S246H/H347A into the beetle luciferase emit luminescence with emission maximum at 626 nm, 88-nm red-shift from the wild-type luciferase. This mutant luciferase is anticipated for application in in vivo multicolor imaging in living samples.
荧光素酶是一种生物发光蛋白，已被用作一种分析工具来可视化细胞内现象。发射红光的荧光素酶因其在哺乳动物组织中的高透射率而特别适用于生物发光成像。然而，现有的发射波长超过600nm的荧光素酶的发光强度较弱，不足以用于成像研究。我们从巴西叩甲（Pyrearinus termitilluminans）中开发了翡翠荧光素酶（Eluc）的突变体，该荧光素酶在甲虫荧光素酶中发出最强的生物发光。我们基于同源建模预测的Eluc结构，依次在荧光素酶中引入了四个氨基酸突变。结果表明，甲虫荧光素酶的四重突变R214K/H241K/S246H/H347A发出的光的发射最大值在626nm，比野生型荧光素酶红移了88nm。这种突变型荧光素酶有望应用于活体样本的体内多色成像。
