Efficient viral delivery system for unnatural amino acid mutagenesis in mammalian cells
Efficient viral delivery system for unnatural amino acid mutagenesis in mammalian cells
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DOI：
10.1073/pnas.1309584110
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发表时间：
2013-07-16
影响因子：
11.1
通讯作者：
Schultz, Peter G.
中科院分区：
文献类型：
--
作者：
Chatterjee, Abhishek;Xiao, Han;Schultz, Peter G.
在这里，我们报道了一种基于杆状病毒的递送系统的发展，该系统能够有效地将非天然氨基酸整合到哺乳动物细胞中的蛋白质中。我们已经利用杆状病毒双链DNA基因组的大载货能力（bbb30 kb）和稳定性，向各种细胞类型传递遗传结合新氨基酸所需的所有成分。这些包括工程tRNA/氨基酰基-tRNA合成酶对和目标基因的无义突变体。杆状病毒在哺乳动物细胞中的转导效率显著提高。将两个多特异性tRNA/氨基酰基-tRNA合成酶对插入到该表达系统中，使多种具有新化学和生物学特性的非天然氨基酸能够特异性地结合到蛋白质中。
Here we report the development of a baculovirus-based delivery system that enables the efficient incorporation of unnatural amino acids into proteins in mammalian cells. We have exploited the large cargo-capacity (> 30 kb) and stability of the double-stranded DNA genome of baculovirus to deliver to a variety of cell types all of the components required to genetically incorporate novel amino acids. These include the engineered tRNA/aminoacyl-tRNA synthetase pair and the nonsense mutant of the target gene. Mammalian cell transduction efficiency of baculovirus was significantly improved by incorporating genetic elements from mammalian viruses. Two polyspecific tRNA/aminoacyl-tRNA synthetase pairs were inserted into this expression system, enabling the site-specific incorporation of a variety of unnatural amino acids with novel chemical and biological properties into proteins.