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通过合理的蛋白质-RNA工程对氨酰-tRNA合成酶进行重新编码用于合成生物学。

Recoding aminoacyl-tRNA synthetases for synthetic biology by rational protein-RNA engineering.

作者信息

Hadd Andrew, Perona John J

机构信息

Department of Biochemistry & Molecular Biology, Oregon Health & Sciences University , 3181 Southwest Sam Jackson Park Road, Portland, Oregon 97239, United States.

出版信息

ACS Chem Biol. 2014 Dec 19;9(12):2761-6. doi: 10.1021/cb5006596. Epub 2014 Oct 31.

DOI:10.1021/cb5006596
PMID:25310879
原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC4273986/
Abstract

We have taken a rational approach to redesigning the amino acid binding and aminoacyl-tRNA pairing specificities of bacterial glutaminyl-tRNA synthetase. The four-stage engineering incorporates generalizable design principles and improves the pairing efficiency of noncognate glutamate with tRNA(Gln) by over 10(5)-fold compared to the wild-type enzyme. Better optimized designs of the protein-RNA complex include substantial reengineering of the globular core region of the tRNA, demonstrating a role for specific tRNA nucleotides in specifying the identity of the genetically encoded amino acid. Principles emerging from this engineering effort open new prospects for combining rational and genetic selection approaches to design novel aminoacyl-tRNA synthetases that ligate noncanonical amino acids onto tRNAs. This will facilitate reconstruction of the cellular translation apparatus for applications in synthetic biology.

摘要

我们采用了一种合理的方法来重新设计细菌谷氨酰胺-tRNA合成酶的氨基酸结合和氨酰-tRNA配对特异性。四阶段工程纳入了可推广的设计原则，与野生型酶相比，非同源谷氨酸与tRNA(Gln)的配对效率提高了10(5)倍以上。蛋白质-RNA复合物的优化设计包括对tRNA球状核心区域的大量重新设计，证明了特定tRNA核苷酸在确定遗传编码氨基酸身份方面的作用。这项工程工作中出现的原则为结合合理和遗传选择方法来设计新型氨酰-tRNA合成酶开辟了新前景，这些酶可将非标准氨基酸连接到tRNA上。这将有助于重建细胞翻译装置，用于合成生物学应用。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/ca7d/4273986/a85c591366b8/cb-2014-006596_0003.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/ca7d/4273986/8c42370ec623/cb-2014-006596_0001.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/ca7d/4273986/f93318c370f1/cb-2014-006596_0002.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/ca7d/4273986/a85c591366b8/cb-2014-006596_0003.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/ca7d/4273986/8c42370ec623/cb-2014-006596_0001.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/ca7d/4273986/f93318c370f1/cb-2014-006596_0002.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/ca7d/4273986/a85c591366b8/cb-2014-006596_0003.jpg

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