An Evolved Aminoacyl-tRNA Synthetase with Atypical Polysubstrate Specificity,
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- 作者：Douglas D. Young ; Travis S. Young ; Michael Jahnz ; Insha Ahmad ; Glen Spraggon ; Peter G. Schultz
- 刊名：Biochemistry
- 出版年：2011
- 出版时间：March 22, 2011
- 年：2011
- 卷：50
- 期：11
- 页码：1894-1900
- 全文大小：890K
- 年卷期：v.50,no.11(March 22, 2011)
- ISSN：1520-4995
文摘
We have employed a rapid fluorescence-based screen to assess the polyspecificity of several aminoacyl-tRNA synthetases (aaRSs) against an array of unnatural amino acids. We discovered that a p-cyanophenylalanine specific aminoacyl-tRNA synthetase (pCNF-RS) has high substrate permissivity for unnatural amino acids, while maintaining its ability to discriminate against the 20 canonical amino acids. This orthogonal pCNF-RS, together with its cognate amber nonsense suppressor tRNA, is able to selectively incorporate 18 unnatural amino acids into proteins, including trifluoroketone-, alkynyl-, and halogen-substituted amino acids. In an attempt to improve our understanding of this polyspecificity, the X-ray crystal structure of the aaRS鈭?i>p-cyanophenylalanine complex was determined. A comparison of this structure with those of other mutant aaRSs showed that both binding site size and other more subtle features control substrate polyspecificity.