An unnatural base pair for incorporating amino acid analogs into proteins
- 1 February 2002
- journal article
- research article
- Published by Springer Nature in Nature Biotechnology
- Vol. 20 (2) , 177-182
- https://doi.org/10.1038/nbt0202-177
Abstract
An unnatural base pair of 2-amino-6-(2-thienyl)purine (denoted by s) and pyridin-2-one (denoted by y) was developed to expand the genetic code. The ribonucleoside triphosphate of y was site-specifically incorporated into RNA, opposite s in a template, by T7 RNA polymerase. This transcription was coupled with translation in an Escherichia coli cell-free system. The yAG codon in the transcribed ras mRNA was recognized by the CUs anticodon of a yeast tyrosine transfer RNA (tRNA) variant, which had been enzymatically aminoacylated with an unnatural amino acid, 3-chlorotyrosine. Site-specific incorporation of 3-chlorotyrosine into the Ras protein was demonstrated by liquid chromatography–mass spectrometry (LC-MS) analysis of the products. This coupled transcription–translation system will permit the efficient synthesis of proteins with a tyrosine analog at the desired position.Keywords
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