Functional replacement of the endogenous tyrosyl-tRNA synthetase-tRNATyr pair by the archaeal tyrosine pair in Escherichia coli for genetic code expansion.
Functional replacement of the endogenous tyrosyl-tRNA synthetase-tRNATyr pair by the archaeal tyrosine pair in Escherichia coli for genetic code expansion.
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DOI：
10.1093/nar/gkq080
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发表时间：
2010-06
影响因子：
14.9
通讯作者：
Sakamoto K
中科院分区：
文献类型：
--
作者：
Iraha F;Oki K;Kobayashi T;Ohno S;Yokogawa T;Nishikawa K;Yokoyama S;Sakamoto K
非天然氨基酸已在活细胞中使用与宿主翻译系统正交的氨酰-tRNA合成酶-tRNA对进行遗传编码。在本研究中，我们用一个与E. coliTyrosyl-tRNA合成酶（TyrRS）-tRNATyr对，利用E. coliTyrRS变体在细胞中用于非天然氨基酸而不干扰酪氨酸掺入。我们证明了E. coliTyrRS-tRNATyr对在功能上可以被詹氏甲烷球菌和酿酒酵母酪氨酸对所取代，它们不与E. coli TyrRS或tRNATyr。然后将内源TyrRS和tRNATyr基因从E.表达古细菌TyrRS-tRNATyr对的大肠杆菌细胞。在该工程菌株中，3-碘-1-酪氨酸和3-叠氮基-1-酪氨酸分别成功地由琥珀密码子编码，使用E.大肠杆菌琥珀抑制子tRNATyr和TyrRS变体，其先前针对3-碘-1-酪氨酸开发并且也被发现识别3-叠氮基-1-酪氨酸。通过X-射线晶体学揭示了3-叠氮基-1-酪氨酸识别的结构基础。目前的工程技术允许E.大肠杆菌TyrRS变异体的非天然氨基酸，以在大肠杆菌中开发。大肠杆菌，用于在真核细胞和细菌细胞中进行遗传密码扩增。
Non-natural amino acids have been genetically encoded in living cells, using aminoacyl-tRNA synthetase–tRNA pairs orthogonal to the host translation system. In the present study, we engineered Escherichia coli cells with a translation system orthogonal to the E. coli tyrosyl-tRNA synthetase (TyrRS)–tRNATyr pair, to use E. coli TyrRS variants for non-natural amino acids in the cells without interfering with tyrosine incorporation. We showed that the E. coli TyrRS–tRNATyr pair can be functionally replaced by the Methanocaldococcus jannaschii and Saccharomyces cerevisiae tyrosine pairs, which do not cross-react with E. coli TyrRS or tRNATyr. The endogenous TyrRS and tRNATyr genes were then removed from the chromosome of the E. coli cells expressing the archaeal TyrRS–tRNATyr pair. In this engineered strain, 3-iodo-l-tyrosine and 3-azido-l-tyrosine were each successfully encoded with the amber codon, using the E. coli amber suppressor tRNATyr and a TyrRS variant, which was previously developed for 3-iodo-l-tyrosine and was also found to recognize 3-azido-l-tyrosine. The structural basis for the 3-azido-l-tyrosine recognition was revealed by X-ray crystallography. The present engineering allows E. coli TyrRS variants for non-natural amino acids to be developed in E. coli, for use in both eukaryotic and bacterial cells for genetic code expansion.
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影响因子：
16.6
作者：
Chen, Peng R.;Groff, Dan;Guo, Jiantao;Ou, Weijia;Cellitti, Susan;Geierstanger, Bernhard H.;Schultz, Peter G.
通讯作者：
Schultz, Peter G.
影响因子：
14.9
作者：
Muyrers, JPP;Zhang, YM;Stewart, AF
通讯作者：
Stewart, AF
DOI：
10.1007/s10969-006-9007-y
发表时间：
2006-03-01
期刊：
Journal of Structural and Functional Genomics
影响因子：
--
作者：
Chumpolkulwong, Namthip;Sakamoto, Kensaku;Yokoyama, Shigeyuki
通讯作者：
Yokoyama, Shigeyuki
DOI：
10.1073/pnas.031488298
发表时间：
2001-02-27
影响因子：
11.1
作者：
Kowal, AK;Köhrer, C;RajBhandary, UL
通讯作者：
RajBhandary, UL
影响因子：
15
作者：
Kwon, Inchan;Tirrell, David A.
通讯作者：
Tirrell, David A.