Click beetle luciferase mutant and near infrared naphthyl-luciferins for improved bioluminescence imaging.
Click beetle luciferase mutant and near infrared naphthyl-luciferins for improved bioluminescence imaging.
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DOI：
10.1038/s41467-017-02542-9
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发表时间：
2018-01-09
影响因子：
16.6
通讯作者：
Mezzanotte L
中科院分区：
文献类型：
--
作者：
Hall MP;Woodroofe CC;Wood MG;Que I;Van't Root M;Ridwan Y;Shi C;Kirkland TA;Encell LP;Wood KV;Löwik C;Mezzanotte L
动物中生物发光成像的灵敏度主要取决于荧光素酶在大于620 nm的波长下发射的光子的量，其中组织穿透性高。这一领域的工作主要是萤火虫荧光素酶及其底物D-荧光素，由于该系统的峰值发射（~ 600 nm），高信噪比，以及D-荧光素在小鼠中的生物分布通常良好。在这里，我们报告了一个密码子优化的突变体的点击甲虫红色荧光素酶，产生实质上更多的光输出比萤火虫荧光素酶时，这两种酶在移植细胞内的黑毛小鼠的皮肤或脑深部进行比较。该突变酶利用两种新的萘酰亚胺底物产生近红外发射（730 nm和743 nm）。稳定的发光信号和近红外发射使得在小鼠中进行深部组织多光谱断层扫描具有前所未有的灵敏度和准确性。需要红移的生物发光发射来提高深部组织成像分辨率。在这里，作者开发了一种点击甲虫红色荧光素酶突变体和两种萘基-荧光素底物，并显示了新的荧光素/荧光素酶配对在小鼠体内进行深层组织多光谱断层扫描的能力。
The sensitivity of bioluminescence imaging in animals is primarily dependent on the amount of photons emitted by the luciferase enzyme at wavelengths greater than 620 nm where tissue penetration is high. This area of work has been dominated by firefly luciferase and its substrate, D-luciferin, due to the system’s peak emission (~ 600 nm), high signal to noise ratio, and generally favorable biodistribution of D-luciferin in mice. Here we report on the development of a codon optimized mutant of click beetle red luciferase that produces substantially more light output than firefly luciferase when the two enzymes are compared in transplanted cells within the skin of black fur mice or in deep brain. The mutant enzyme utilizes two new naphthyl-luciferin substrates to produce near infrared emission (730 nm and 743 nm). The stable luminescence signal and near infrared emission enable unprecedented sensitivity and accuracy for performing deep tissue multispectral tomography in mice. Red-shifted bioluminescence emission is needed to improve deep tissue imaging resolution. Here, the authors develop a click beetle red luciferase mutant and two naphthyl-luciferin substrates, and show the ability of the new luciferin/luciferase pairing for deep tissue multispectral tomography in mice.
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